IkkikiI  Fi'bruury  7,  liOi 

U.  S.  DEPARTMENT  OF  AGRICULTURE, 

BUREAU  OF  ANIMAL  INDUSTRY.— BULLETIN  118. 
A.  D.  MELVIN,  Chief  op  Bureau. 


S  =  ULTURAL  STUDIES  OF  SPECIES 
OF  PENICILLIUM. 


BY 


CHARLES  THOM,  Ph.  D., 

Mycologist  in  Cheese  Investigations,  Dairy  Division. 


JAN  4    1943 
^^  il  fe  R  A 


WASHINGTON: 

GOVERNMENT   PRINTING   OFFICE. 

1910. 


Issued  February  7, 1910. 

U.  S.  DEPARTMENT  OF  AGRICULTURE, 

BUREAU  OF  ANIMAL  INDUSTRY.— Bulletin  118. 

A.  D.  MELVIN,  Chief  of  Bureau. 


CULTURAL  STUDIES  OF  SPECIES 
OF  PENICILLIUM. 


BY 


CHARLES  THOM,  Ph.  D., 

Mycologist  in  Cheese  Investigations,  Dairy  Division. 


WASHINGTON: 

GOVERNMENT   PRINTING  OFFICE. 

1910. 


THE  BUREAU  OF  ANIMAL  INDUSTRY. 


Chief:  A.  D.  Melvin. 
Assistant  Chief:  A.  M.  Farrington. 
Chief  Clerk:  Charles  C.  Carroll. 
Biochemic  Division:  M.  Dorset,  chief. 
Dairy  Division:  B.  H.  Rawl,  chief. 

Inspection  Division:  Rice  P.  Steddom,  chief;  Morris  Wooden,  R.  A.  Ramsay, 
and  Albert  E.  Behnke,  associate  chiefs. 
Pathological  Division:  John  R.  Mohler,  chief. 
Quarantine  Division:  Richard  W.  Hickman,  chief. 
Zoological  Division:  B.  H.  Ransom,  chief. 
Experiment  Station:  E.  C.  Schroeder,  superintendent. 
Animal  Husbandry  Division:  George  M.  Rommel,  chief. 
Editor:  James  M.  Pickens. 


LETTER  OF  TRANSMITTAL. 


.     U.  S.  Department  of  Agriculture, 

Bureau  of  Animal  Industry, 

Washington,  D.  C,  July  12,  1909. 

Sir:  I  have  the  honor  to  transmit  herewith  and  to  recommend  for 
pubhcation  as  a  bulletin  of  this  Bureau  a  manuscript  entitled  ''Cul- 
tural Studies  of  Species  of  Penicillium,"  by  Dr.  Charles  Thom,  my- 
cologist in  the  cooperative  soft-cheese  investigations  carried  on  at 
Storrs,  Conn.,  by  the  Storrs  Agricultural  Experiment  Station  and 
the  Dairy  Division  of  this  Bureau.  A  previous  paper  of  this  author 
deaUng  with  certain  species  of  Penidllium  which  are  characteristic 
factors  in  the  ripening  of  Camembert  and  Roquefort  cheese  has  been 
pubhshed  as  Bulletin  82  of  this  Bureau.  The  investigations  con- 
nected with  the  study  of  the  organisms  referred  to  necessitated  the 
culture  and  comparison  of  many  other  species  and  an  examination  of 
the  nomenclature  for  the  whole  genus  Penicillium.  Considerable  con- 
fusion was  found  to  exist  regarding  the  identification  of  the  various 
species,  hence  it  seemed  important  that  a  comprehensive  study  from 
cultural  data  of  all  obtainable  species  should  be  undertaken. 

Acknowledgment  is  made  of  suggestions  and  advice  by  Dr.  Erwin 
F.  Smith,  of  the  Bureau  of  Plant  Industry  of  this  Department,  be- 
sides the  persons  whose  names  are  given  in  the  text. 

The  illustrations,  with  one  exception,  are  from  original  drawings 
by  the  author. 

Respectfully,  A.  D.  Melvin, 

Chief  of  Bureau. 
Hon.  James  Wilson, 

Secretary  of  Agriculture. 


Digitized  by  the  Internet  Archive 

in  2007  with  funding  from 

IVIicrosoft  Corporation 


http://www.archive.org/details/culturalstudiesoOOthomiala 


CONTENTS. 


Page. 

Introduction , 9 

The  basis  for  specific  characterization 10 

Necessity  for  describing  culture  media  and  temperature  conditions 14 

Habit,  structure,  and  appearance  of  colonies 15 

Physiological  effects  upon  media 19 

Culture  media 22 

Nomenclature 23 

The  generic  name 23 

Nomenclatureof  species 23 

The  type  species 25 

Explanation  of  drawings 27 

Penicillium  expansum  Link 27 

Penicillium  italicum  Wehmer 29 

Penicillium  digitatum  Saccardo 31 

Penicillium  roque/orti  Thom 34 

Penicillium  purpurogenum  O.  StoU 36 

Penicillium  pinophilum  Hedgcock  (nomen  novum) 37 

Penidlliiim  rubrum  O.  Stoll 39 

Penicillium  luteum  Zukal 39 

Penicillium  duclav^ci  Delacroix 42 

Penicillium  claviforme  Bainier 43 

Penicillium  granulatum  Bainier 44 

Penicillium  brevicaule  Saccardo 45 

Penicillium  brevicaule  Saccardo,  var.  album  Thom,  n.  var 47 

Penicillium  brevicaule  Saccardo,  var.  glabrum  Thom,  n.  var 48 

Penicillium  roseum  Link  (?) 49 

Penicillium  cam^mberti  Thom 50 

Penicillium  camemberti  var.  rogeri  Thom,  n.  var 52 

Penixillium  bi/orme  n.  sp 54 

Penicillium,  commune  n.  sp 56 

Penicillium  No.  22 58 

Penicillium  chrysogenum  n.  sp - 58 

Penicillium  rugulosum  n.  sp 60 

Penicillium,  citrinum  n.  sp 61 

Penicillium  No.  37 63 

Penicillium  No.  12 64 

Penicillium  alramenlosum  n.  sp 65 

Penicillium  No.  24 " 66 

Penicillium  sloloniferum  n.  sp 68 

Penicillium funicuhsum  n.  sp 69 

Penicillium  decumbens  n.  sp 71 

Penicillium  divaricatum  n.  ap 72 

Penicillium  lilacinum  n.  sp 73 

5 


6  CONTENTS. 

Page. 

Penidllium,  intricatum  n.  sp 75 

Penicillium  spiniilosuin  n.  hj) 76 

Penicillium  No.  28 77 

Species  forming  pink  sclerotia 78 

Penicillium  No.  29 78 

Penicillium  No.  30 79 

Penicillium  No.  31 80 

Penicillium  No.  32 80 

Comparative  cultural  data 81 

Cultures  in  distilled  water 82 

Agar-agar  as  a  source  of  food 83 

Agar-agar  as  a  source  of  carbon 83 

Various  sources  of  carbon 83 

Cultures  in  Raulin's  fluid  and  in  Cohn's  solution 86 

Color  in  conidial  areas 87 

Effect  of  concentrated  media 88 

The  grouping  of  species 89 

Odors 90 

Anaerobic  cultures  (with  carbon  dioxid) 90 

Incubation  te.sts 91 

Summary  of  data  from  comparative  culture 94 

Keys  to  cultural  identification  of  species 94 

Key  1 . — Analysis  of  species  in  cultures  upon  gelatin  and  agar 95 

Key  2. — Species  determinable  from  substrata 97 

Tabular  statements 98 

Table  1 . — Tabulation  of  salient  characters  of  species 98 

Table  2. — Gelatin  and  color  reactions 100 

Table  3. — Comparative  cultures 101 

Table  4. — Comparative  cultures  in  synthetic  fluid  (Dox's) 102 

Table  5. — Decimal  summary  of  comparative  cultures  in  synthetic  fluid 

(Dox's) 104 

Table  6. — Incubation  experiments 105 

References  to  literature lOG 

Index  to  species 109 


ILLUSTRATIONS. 


Page. 

Fig.  1 .  Penicillium  expansum  Link 28 

2.  Penicillium  italicum  Wehmc^r 30 

3.  Penicillium  digitatum  Saccardo 33 

4.  Penicillium  roqueforli  Thorn 35 

5.  Penicillium  purpurogenum  O.  Stoll 37 

6.  Penicillium  pinophilum  Hedgcock 38 

7.  Penicillium  rubrum  O.  Stoll 40 

8.  Penicillium  luteum  Ziikal 41 

9.  Penicillium  duclauxi  Delacroix 42 

10.  Penicillium  clavi/orme  Bainicr 43 

n .  Penicillium  granulalum  Bainier 45 

12.  Penicillium  brevicaule  Saccardo 46 

13.  Penicillium  brevicaule,  var.  album 47 

14.  Penicillium  brevicaule,  var.  glibrum 48 

15.  Penicillium  roseum  Link  (?) 49 

16.  Penicillium  camemberti  Thorn 51 

17.  Penicillium  camemberti,  var.  rogeri 53 

18.  Penicillium  bi/orme 55 

19.  Penicillium.  commune 57 

20.  Penicillium  chrysogenum 59 

21.  Penicillium  rugulosum 61 

22.  Penicillium  citrinum 62 

23.  Penicillium  No.  37 64 

24.  Penicillium  atramentosuin 66 

25.  Penicillium  No.  24 67 

26.  Penicillium  stoloniferum 68 

27.  Penicillium funiculosum 70 

28.  Penicillium  decumbens 71 

29.  Penicillium  divaricalum 73 

30.  Penicillium  lilacinum 74 

31 .  Penicillium  intracntum 75 

32.  Penicillium  spinulosum 76 

33.  Penicillium  No.  28 77 

34.  Penicillium  No.  29 79 

35.  Penicillium  No.  31 , 80 

36.  Penicillium  No.  32 81 

7 


CULTURAL  STUDIES  OF  SPECIES  OF  PENICILLIUM. 


INTRODUCTION. 

In  a  previous  paper  ^^'^  two  species  of  PeniciUium  were  shown  to 
secrete  the  proteolytic  enzyms  which  ripen  certain  varieties  of 
cheese.  The  effort  to  identify  these  organisms  necessitated  the 
culture  and  comparison  of  numerous  other  species  and  a  study  of 
the  literature  of  nomenclature  for  the  whole  of  the  genus  PeniciUium. 
The  difficulties  encountered  in  deciding  whether  to  discuss  these 
forms  under  old  names  or  to  describe  them  anew  from  cultural  data 
led  to  the  extension  of  this  study  beyond  the  forms  occurring  in  a 
dairy  investigation  so  as  to  include  any  obtainable  species,  and 
especially  all  species  whose  identification  under  published  names 
could  be  established. 

Many  recent  studies  have  linked  particular  chemical  and  physio- 
logical activities  with  the  presence  of  particular  species  of  fungi. 
When  such  data  relate  to  known  species — identifiable  species — our 
knowledge  of  the  metabolism  of  these  forms  has  been  greatly  in- 
creased. When,  as  has  often  happened,  the  generic  name  alone  is 
given  in  a  group  so  diverse  in  its  activities  as  PeniciUium,  such  data 
only  add  to  the  confusion.  Whether  the  observations  apply  to  all 
of  the  species  of  the  genus  or  to  a  single  unnamed  species  is  left  un- 
settled. To  give  real  utility  to  such  work,  some  one  must  test  the 
applicability  of  the  data  to  each  species  under  consideration.  On 
account  of  their  ease  of  cultivation  and  the  notion  that  there  was  a 
single  common  green  species  legitimately  named  P.  glaucum,  species 
of  PeniciUium  have  formed  the  subjects  of  many  such  investigations. 
In  very  few  of  these  cases  are  sufficient  data  regarding  morphology 
given  to  warrant  even  a  guess  as  to  the  species  used.  An  examination 
of  the  present  status  of  specific  nomenclature  in  the  genus  will  there- 
fore furnish  a  sounder  basis  for  further  studies  of  their  activities. 

This  paper  represents  cultural  work  which  has  continued  more 
than  four  years  and  includes  those  species  for  which  the  data  obtained 
seem  abundantly  to  justify  the  characterization  oiTered.  Some  of 
these  forms  have  been  cultivated  for  the  whole  period,  others  for  a 
less  time.     No  claim  to  monographic  completeness  can  be  made.     In 

oThe  figures  refer  to  the  list  of  literature  at  end  of  bulletin. 


10  CULTURAL  STUDIES  OP  SPECIES  OF  PENICILLIUM. 

collecting  these  forms,  many  of  them  have  been  isolated  in  this 
laboratory  from  dairy  products  and  from  fruits,  or  collected  in  the 
field  by  the  writer.  One  series  of  forms  was  purchased  from  Krai,  of 
Prague.  Several  species  have  been  contributed  by  or  verified  by 
those  who  described  them — two  by  Dr.  G.  Bainier,  two  by  Dr.  C. 
Wehmer,  one  by  Dr.  G.  Delacroix,  and  one  by  Dr.  G.  G.  Hedgcock. 
Many  correspondents  have  sent  cultures  for  examination  or  study. 

THE  BASIS  FOR  SPECIFIC  CHARACTERIZATION. 

The  available  sources  of  identification  of  these  species  will  be 
discussed  first.  By  the  kindness  of  Professor  Thaxter  the  exsiccati 
of  the  genus  in  the  herbarium  of  Harvard  University  were  examined. 
Cultures  of  certain  species  were  tried  but  no  living  spores  were 
found.  No  specimen  was  found  in  such  condition  as  to  be  used  to 
identify  material  by  comparison.  Similar  courtesy  was  extended 
for  the  examination  of  specimens  in  the  herbarium  of  Kew  Gardens 
and  at  the  University  of  Berlin.  In  no  case  was  material  found  by 
which  cultural  material  could  be  identified  or  identification  verified 
by  comparison.  The  plant  bodies  are  too  evanescent  to  retain  the 
criteria  necessary  for  identification  for  any  great  length  of  time,  as 
ordinarily  preserved.  In  most  cases  after  a  few  years  of  handling 
the  specimens  were  found  reduced  to  powder.  Wehmer  ^^  has  noted 
for  certain  species  of  this  genus  that  the  conidia  do  not  retain  their 
power  to  germinate  beyond  a  very  few  years.  Specimens  as  formerly 
prepared  therefore  become  useless  for  the  identification  of  species 
by  the  method  of  types.  The  method  described  by  Hedgcock*  for 
preserving  specimens  has  thus  far  been  applied  only  to  one  species 
of  Penicillium  and  entirely  too  recently  to  test  its  permanent  value. 
Aside,  therefore,  from  certain  species  which  will  be  discussed  later, 
it  has  been  impossible  to  determine  material  belonging  to  this  genus 
from  herbarium  specimens. 

There  remain  two  methods  of  identification:  (1)  The  identification 
of  material  or  cultures  by  the  authors  of  descriptions;  (2)  identifica- 
tion by  critical  study  of  the  descriptions  and  illustrations  published. 

Regarding  the  first  method,  fortunately  some  of  the  more  recent 
authors  have  either  preserved  their  organisms  in  culture,  as  is  done 
at  the  Ecole  de  Pharmacie  in  Paris  with  the  cultures  of  Bainier,  or 
placed  them  in  one  other  of  the  distributing  laboratories  where 
such  organisms  are  maintained  in  continuous  culture.  But  the  earlier 
authors  are  dead  and  have  left  only  their  published  descriptions  and 
figures  as  a  means  of  determining  what  organisms  they  studied. 

Turning  to  the  critical  comparison  of  material  with  published 
descriptions,  we  find  many  difficulties.  These  descriptions  give  in 
meager  outline  observations  of  mold  masses  showing  penicillate  coni- 
dial  fructifications,  as  found  in  nature  upon  substrata  more  or  less 


BASIS  FOR   SPECIFIC   CHARACTERIZATION.  11 

accurately  specified.  No  cultures  were  made.  The  original  masses 
are  assumed  to  be  comprised  each  of  a  single  species.  Parasitism  or 
selective  saprophytism  is  assumed,  but  the  substratum  is  rarely 
designated  with  suflicient  care  to  make  a  duplication  of  the  original 
culture  possible.  Hence  the  characters  would  include  whole  series 
of  forms  whose  differences  are  marked.  No  account  is  taken  of  their 
omnivorous  nature  nor  of  the  marked  variations  introduced  in  ap- 
pearance by  changed  conditions.  The  method  of  types  and  the  method 
of  substrata  as  represented  by  the  herbarium  material  and  the  pub- 
lished descriptions  we  have  are,  therefore,  not  sufficient  for  the 
identification  of  species  of  Penicillium. 

Although  we  abandon  the  "method  of  substrata"  as  the  sole  basis 
of  description,  the  force  of  natural  selection  as  shown  by  the  distri- 
bution of  certain  of  these  species  in  nature  is  a  most  valuable  acces- 
sory. Some  species  occur  so  constantly  upon  particular  substrata 
and  under  particular  conditions  as  to  simplify  their  identification 
greatly.  Examples  of  this  are  P.  italicum  Wehmer  and  P.  digitatum 
Saccardo,  as  they  are  found  upon  citrus  fruits.  Unfortunately  very 
few  species  restrict  themselves  to  particular  substrata,  so  that  except 
for  some  few  species  and  a  very  few  media,  the  occurrence  of  a 
Penicillium  in  any  given  situation  is  but  slight  evidence  for  its 
identification.  The  constant  occurrence  of  species  of  Penicillium 
in  the  laboratory,  in  connection  with  foods,  and  in  factory  processes, 
such  as  cheese-ripening,  all  point  to  controlled  culture  as  the  proper 
source  of  diagnostic  characters. 

If  we  look  to  cultural  study  "■  for  our  conception  of  species  we  have 
two  methods  of  procedure:  (1)  The  exhaustive  study  of  the  limits  of 
variability  for  each  species;  (2)  the  comparative  study  of  numerous 
species  under  arbitrarily  chosen  conditions  uniformly  maintained, 

Tlie  first  is  the  best  method  known  for  gaining  complete  knowledge 
of  single  species,  but  it  is  too  cumbrous  for  taxonomic  purposes. 
Physiological  and  chemical  studies  have  commonly  been  restricted 
to  particular  classes  of  reactions  for  single  species  or  groups  of  species. 
These  have  contributed  much  to  our  knowledge  of  fungous  varia- 
bility, but  too  often  give  no  hint,  except  the  vague  nomenclature 
used,  upon  which  to  judge  which  species  were  actually  studied.  In 
experimental  cultures  changes  in  the  chemical  nature  of  the  medium 
.or  in  the  conditions,  or  both,  have  been  found  to  produce  great 
changes  in  the  morphology  of  the  fungi  studied.  With  the  exception 
of  a  few  fundamental   group    or   generic  characters,  nearly  every 

oWeidemann,'®  in  a  recent  paper,  has  described  several  new  species  of  this  genus 
in  their  relation  to  culture  media.  In  this  he  has  followed  the  bacteriologists  further 
than  the  writer  has  thought  necessary  by  giving  formulas  for  a  considerable  number 
of  media  and  detailed  notes  as  to  reactions  upon  such  media,  instead  of  the  more 
formal  descriptions  attempted  here. 


12  CULTURAL  STUDIES  OF  SPECIES  OF  PENICILLIUM. 

attribute  used  in  specific  description  has  been  shown  to  be  a  reaction 
to  environment,  hence  changeable  with  such  environment  (for  some 
species  at  least).  Exhaustive  study  of  all  the  species  would  be 
endless. 

The  alternative  is  to  select  certain  media  and  a  particular  set  of 
conditions,  then  to  cultivate  all  the  organisms  under  investigation 
in  a  uniform  manner  and  to  base  distinctions  of  species  upon  differ- 
ences in  the  reactions  obtained,  and  upon  the  differing  characters  of 
the  several  species,  in  this  common  environment. 

To  test  the  reliability  of  such  data,  species  obtained  in  the  dairy 
laboratories  were  first  studied  carefully  upon  the  peptone-milk-sugar 
gelatin  described  by  Conn*  and  upon  potato  agar  as  described  in 
our  previous  bulletin  (Thom,"  p.  7).  From  these  cultures  trans- 
fers have  been  made  to  media  of  very  different  composition — Cohn's 
solution,  Raulin's  fluid,  milk  in  various  forms,  synthetic  fluids  pre- 
senting different  sources  of  carbon — always  bringing  cultures  back 
to  the  original  media.  Many  species  differ  so  materially  in  gross 
characters  when  grown  upon  these  different  media  that  successive 
cultures,  if  not  kiown  to  be  pure  transfers,  might  be  supposed  to 
be  different  species;  but  when  returned  to  the  original  media  and 
conditions  these  forms  have  immediately  produced  the  characters 
and  reactions  first  found,  with  a  large  degree  of  uniformity.  Abso- 
lutely uniform  reactions  are  not  to  be  expected  from  living  organ- 
isms, at  any  rate  under  our  imperfect  control  of  working  conditions ; 
but  when  such  reactions  are  definitely  recognizable  as  essentially 
the  same,  the  result  may  be  judged  as  satisfactory.  It  is  even  more 
confusing  to  find  that  two  or  more  species  may  react  very  similarly 
upon  a  particular  substratum.  A  transfer  of  these  organisms  to  a 
medium  of  markedly  different  composition  brings  out  the  contrasting 
characters,  however.  The  desirability  of  recording  the  widest  pos- 
sible distinctions  in  such  descriptive  work  makes  necessary  the  use 
of  media  differing  in  composition  as  much  as  practicable,  in  such 
comparative  cultures. 

It  is  worthy  of  note  that  the  species  P.  roqueforti  and  P.  camemherti, 
essential  to  the  cheese  industry,  have  been  isolated  repeatedly  from 
cheeses  of  widely  different  origin.  The  Roquefort  species  has  been 
obtained  from  laboratories  not  concerned  with  Roquefort  cheese 
studies,  from  ensilage,  and  from  other  substances.  The  same  char- 
acters have  been  found  in  cultures  of  these  two  species  from  these 
variable  sources  under  conditions  in  which  the  possibility  of  close 
genetic  connection  between  cultures  is  thus  remote.  There  seems  to 
be  little  possibility  of  question  that  these  species  at  least  are  well- 
fixed  conidia-bearing  forms,  not  cultural  varieties  of  other  species. 
The  question  at  issue  was  not  whether  or  how  variations  could  be 
produced,  but  whether  a  particular  variation  is  constantly  produced 


BASIS  FOR  SPECIFIC   CHARACTERIZATION.  13 

by  a  species  in  a  particular  environment.  The  correlative  question 
whether  the  characters  of  a  given  species  of  mold  can  be  perma- 
nently changed  by  passing  through  a  series  of  cultures  upon  different 
media  is  involved  in  the  same  investigation.  It  is  asserted  by  some 
workers  that  the  physiological  reactions  of  molds  (if  not  also  the 
morphology)  can  be  changed,  and  that  such  changes  persist  after 
the  return  of  the  species  to  the  original  environment.  So  far  as  this 
investigation  has  gone  such  a  view  is  certainly  not  supported  by  the 
conduct  of  the  species  of  Penicillium  which  have  been  studied.  In 
those  species  most  thoroughly  studied  both  the  physiological  and 
morphological  reactions  have  appeared  to  be  very  reliable.  A  sum- 
mary of  these  observations  follows: 

1.  The  same  species  may  differ  greatly  in  morphology  and  physi- 
ological reactions  when  grown  upon  different  media. 

2.  Two  species  closely  similar,  when  grown  parallel  in  one  en- 
vironment, may  differ  characteristically  when  transferred  to  a  differ- 
ent medium  or  a  different  set  of  conditions. 

3.  With  these  species  the  repetition  of  culture  under  particular 
conditions  produces  fairly  constant  morphology  and  reactions. 

Forms  arising  in  this  way  have  been  designated  as  ecads  by  Clem- 
ents^ in  a  recent  discussion  of  the  "Aspects  of  the  species  question." 
This  name  is  used  preferably  for  forms  whose  origin  is  kno\vn  either 
because  produced  by  cultivation  or  under  such  carefully  determined 
natural  conditions  as  admit  of  full  description.  With  species  of 
this  genus  in  which  part  at  least  of  the  morphological  characters  of 
every  culture  are  definitely  attributable  to  the  conditions  and  to  the 
chemical  character  of  the  substratum,  practically  all  kno\vTi  forms 
would  therefore  be  properly  designated  as  such  ecads.  If  described 
as  ecads,  however,  each  description  must  be  liriiited  strictly  to  the 
data  obtainable  upon  a  single  medium,  whereas  by  writing  into  the 
description  the  reference  to  media  it  has  been  found  possible  to  in- 
clude a  much  more  complete  set  of  characters  than  would  readily  be 
worked  out  from  a  single  set  of  conditions. 

These  observations  lead  to  the  conclusion  that  the  cultural  descrip- 
'  tion  of  species  of  molds  demands  the  recognition  of  the  points  noted 
below: 

1.  The  culture  media  and  conditions  must  be  described  so  fully  as 
to  make  the  repetition  of  the  culture  upon  the  same  medium  and  under 
approximately  the  same  conditions  easily  possible  anywhere. 

2.  The  habit,  structure,  and  appearance  of  the  colony  must  be 
given  as  it  develops  upon  at  least  two  standard  media  of  decidedly 
different  composition. 

3.  The  physiological  effects  of  the  colony  upon  these  media  should 
be  noted. 


14  CULTURAL  STUDIES   OF   SPECIES  OF  PENICILLIUM. 

4.  Full  drawings  or  photograplis  should  show  habit  as  well  as  micro- 
scopic details  of  cells  and  cell  relations. 

5.  Other  morphological  or  physiological  data  obtainable  should 
be  given  as  accessory  information.  Very  striking  characters  are 
often  found  under  accidental  or  unique  conditions  which  immedi- 
ately differentiate  particular  species.  Some  of  these  characters  come 
up  in  the  ordinary  course  of  cultural  study,  others  are  found  under 
accidental  conditions  but  could  rarely  occur  in  laboratory  routine. 

It  is  clear  that  such  descriptions  can  only  result  from  repeated  cul- 
ture of  a  species  imder  constant  observation.  If  the  range  of  such 
culture  has  been  wide,  it  will  bring  out  the  most  striking  characters 
and  thus  reduce  the  number  of  minute  distinctions  necessary. 

NECESSITY    FOR   DESCRIBING   CULTURE    MEDIA   AND   TEMPERATURE 

CONDITIONS. 

In  a  previous  paper  *^  the  characters  available  from  cultural  studies 
of  species  of  Penicillium  were  discussed.  The  series  of  observations 
have  been  greatly  extended  in  the  three  years  intervening.  These 
characters  may  therefore  be  profitably  reviewed,  following  the  sum- 
mary just  given. 

Culture  medium. — The  composition  of  the  substratum  is  shown  in 
this  paper  to  affect  the  character  of  the  colonies  grown  upon  it  to 
such  a  degree  as  to  make  the  exact  description  of  the  medium  essen- 
tial. Examples  of  these  effects  may  be  cited  to  show  how  conspicu- 
ous these  differences  may  be.  In  a  medium  free  from  certain  sugars 
P.  duclauxi  produces  upon  the  surface  of  the  medium  very  short  coni- 
diophores  with  conidial  fructifications,  whereas  when  such  a  sugar 
is  added  numerous  coremia  are  formed,  which  in  well-nourished  colo- 
nies often  become  10  mm.  in  height.  This  species  also  produces  a 
rich  purple  color  in  certain  media,  but  not  in  others.  Hedgcock  has 
noted  that  P.  aureum  Corda  (as  distributed  by  him)  produces  colo- 
nies orange-red  upon  alkaline  media,  but  lemon-yellow  in  acid  media. 
P.  digitatum  Sacc.  grows  sparingly,  if  at  all,  in  media  offering 
nitrogen  only  as  nitrates,  whereas  many  other  species  grow  equally 
well  from  nitrates  and  organic  nitrogen.  One  species  is  included 
which  produces  feeble  gray  or  brownish  cultures  when  carbon  is  pre- 
sented from  gelatin,  starch,  or  lactose,  but  becomes  a  clear  green 
when  cane  sugar  is  added. 

A  culture  medium  must  offer  not  only  the  proper  chemical  ele- 
ments for  fungous  growth,  but  must  offer  these  in  assimilable  form. 
Different  species  of  Penicillium  make  quite  different  demands  as  to 
the  form  of  food  substances  required,  hence  a  description  must  specify 
the  substratum  accurately  enough  to  enable  the  duplication  of  cul- 
ture conditions.  If,  however,  the  proper  substances  in  the  proper 
form  are  offered,  changed  percentages  in  the  concentrations  of  these 


HABIT,  STRUCTURE,  AND  APPEARANCE.  15 

substances  have  been  found  to  affect  the  growth  of  cultures  more 
slowly.  Most  species  grow  normally  in  extremely  dilute  solutions, 
but  continue  to  grow  well  or  even  richly  until  the  solutions  reach  con- 
centrations of  considerable  osmotic  pressure. 

Temperature. — The  species  studied  are  mostly  saprophytes  of  wide 
distribution.  They  are  therefore  able  to  thrive  within  a  consider- 
able range  of  temperature.  Comparatively  few  of  the  species  tested 
grew  normally  at  blood  heat  (37°  C).  At  this  maximum  many  of 
them  either  failed  to  grow  or  were  actually  killed.  Within  the  range 
of  12°  to  30°  C.  the  rapidity  of  development  in  most  species,  as 
indicated  by  the  production  of  colored  fruit,  increased  with  the  rise 
in  temperature.  The  lower  temperatures  affect  the  rate  of  fruit  for- 
mation without,  as  a  rule,  preventing  such  development.  Cultures 
held  at  10°  to  20°  C.  reached  a  development  fully  equal  to  those  held 
at  higher  temperature,  only  in  longer  periods. 

HABIT,  STRUCTURE,  AND  APPEARANCE  OF  COLONIES. 

Many  series  of  comparative  cultures  indicate  that  any  colony  of  a 
particular  species  will  reproduce  the  same  characters  whenever 
grown  upon  a  particular  medium  under  particular  conditions.  The 
differences  between  many  species  of  Penicillium  are  so  striking  to  the 
eye  as  to  enable  one  familiar  with  them  to  identify  the  several  species 
immediately.  These  same  differences  are,  however,  so  difficult  to 
define,  and  in  many  cases  so  transitory,  as  to  render  their  expression 
in  form  to  insure  recognition  very  difficult.  Recognition  of  species 
depends  at  best  upon  a  series  of  observations  of  these  characters 
throughout  one,  and  usually  more  than  one,  generation  upon  two  or 
more  substrata.     A  discussion  of  these  characters  follows: 

Fruiting  period. — In  comparing  cultures  of  related  forms  it  is 
found  that  the  time  necessary  for  development  from  the  spore  to  the 
production  of  ripe  conidia  differs  for  the  different  species,  but  that 
periods  in  the  different  species  bear  a  comparatively  constant  rela- 
tion to  each  other  when  all  are  grown  under  the  same  conditions. 
Similarly  the  length  of  the  growing  and  fruiting  period  varies  in  the 
several  species.  In  some  species  the  mycelium  produces  new  coni- 
diophores  and  masses  of  conidia  among  or  overgrowing  the  old  for  a 
considerable  length  of  time.  In  some  a  secondary  growth  of  white 
or  even  colored  hyphse  often  overgrows  the  fruiting  area.  In  others 
the  chains  of  conidia  once  started  continue  to  increase  in  length  for 
several  weeks,  until  the  conidial  fructification  becomes  a  mass  1  mm. 
or  even  more  in  length.  In  still  others  but  a  single  crop  of  conidia 
is*developed  and  the  mycelium  apparently  dies  or  is  totally  exhausted 
in  a  few  days.  In  some  species  the  center  of  the  colony  matures  and 
dies  while  the  margin  continues  to  grow  and  produce  new  conidio- 
phores  for  some  time.    The  habit  of  the  species  in  the  production  of 


16  CULTURAL  STUDIES   OF   SPECIES  OF  PENICILLIUM. 

coiiidial  fruit  is  usually  a  well-marked  character  and  is  often  found 
very  useful  in  the  separation  of  forms  found  growing  together. 

Color. — Color  and  color  changes  are  difficult  to  describe  on  account 
of  the  deficiencies  in  the  standards  of  color,  but  they  form  the  first 
character  noticed.  The  variety  of  greens,  blue  greens,  gray  greens, 
yellow  greens,  and  shades  of  brown  in  the  genus  Penidllium  baffles 
one  seeking  descriptive  terms.  The  shades  of  color  peculiar  to  each 
species  under  oft-repeated  conditions  are  easily  recognized  and  are 
quite  reliable.  The  alterations  in  the  color  of  spores  due  to  changing 
the  composition  of  the  medium,  as  shown  in  the  recent  work  of  StolP* 
and  in  this  paper  for  species  of  this  genus,  and  by  others  (Milburn,*^ 
Bessey  0  for  other  genera,  emphasize  again  the  necessity  of  uniformity 
in  and  careful  description  of  the  culture  medium.  But  in  spite  of 
the  admitted  difficulties  in  color  description,  the  careful  observation 
and  record  of  the  color  of  the  colony  at  every  stage  of  its  development 
is  very  necessary  to  identification.  This  is  complicated  by  the 
changes  which  occur  at  different  ages  of  the  colonies,  so  that  it  would 
be  easily  possible  to  place  certain  species  in  at  least  two  of  the  color 
groups  as  designated  by  the  older  authors  if  we  simply  make  our 
observations  a  few  days  apart. 

The  color  of  the  mass  of  mycelium  as  observed  from  below  ("re- 
verse" as  designated  by  Dierckx^)  in  cultures  gives  striking  con- 
trasts. This  must  not  be  confused  with  discoloration  of  the  sub- 
stratum which  may  or  may  not  be  produced  by  the  same  species  in  a 
given  medium.  The  mycelium  itself  viewed  in  reverse  has  character- 
istic colors  in  certain  species  which  are  useful  in  diagnosis  and  are 
entirely  independent  of  discolorations  of  the  substratum.  A  colony 
colorless  itself  may  color  the  medium  brightly,  while  a  colony  bright 
colored  itself  may  make  no  change  in  the  color  of  the  medium. 

Surface. — As  a  convenient  term  "surface"  may  be  used  to  desig- 
nate the  general  appearance  or  the  texture  of  the  aerial  portion  of 
the  colony.  Perhaps  the  word  "habit"  would  be  in  some  measure 
more  accurate,  but  that  would  apply ^  also  to  the  submerged  myce- 
lium. Comparison  of  the  surface  appearances  of  many  cultures 
shows  that  this  is  one  of  the  most  stable  characters  when  the  same 
medium  and  conditions  are  used.  For  species  of  Penicillium  and 
allied  genera  two  types  of  surface  will  include  most  of  the  species 
met  with. 

In  the  one  type  all  or  a  large  majority  of  the  conidiophores  in  the 
rapidly  growing  colony  arise  directly  from  strictly  vegetative  hyphss 
which  may  be  submerged  in  the  substratum  or  lie  upon  its  surface 
or  be  alternately  prostrate  and  submerged.  Each  conidiophore 
stands  separately,  therefore,  and  usually  all  are  found  to  be  so  nearly 
of  the  same  length  that  the  sui-face  appears  to  be  velvety.  vSuch  a 
surface  may  be  called  either  velvety  or  "strict."  A  strict  surface 
may  be  called  "closely  strict"  when  the  conidiophores  are  so  short 


SURFACE  AND   MARGIN   OF   COLONIES.  17 

that  the  conidial  fructifications  are  barely  raised  above  the  surface, 
or  it  may  be  loose  or  lax  if  longer  coniciiophores  give  a  deeper 
velvety  appearance. 

In  the  other  type  all  or  most  of  the  conidiophores  are  lateral 
branches  of  definitely  aerial  hypha?.  These  hyphse  and  conidiophores 
form  felted  masses  or  loose  networks  of  aerial  mycelium  for  which 
the  term  "floccose"  is  descriptive. 

Margin. — In  seeking  the  origin  of  all  structures  direct  observation 
of  the  margin  of  the  young  and  growing  colony  is  essential.  Such 
observation  determines  how  the  fungus  spreads  in  the  substratum, 
the  septation  and  measurement  of  hypha),  and  the  origin,  order  of  de- 
velopment, and  relative  positions  of  aerial  structures.  As  the  colony 
matures  growth  ceases,  ripe  conidial  areas  extend  to  the  very  margin, 
the  masses  of  conidia  often  change  color  and  fall  apart,  and  the 
conitlia-bearing  branches  may  curl  up  or  drop  off.  Some  species  seem 
to  inliibit  their  own  further  growth  after  a  short  period,  wliile  in  other 
cases  they  dry  up  the  culture  medium  by  transpiring  water.  Many 
things  thus  contribute  to  render  the  old  colony  an  unintelligible  mass 
of  spores  and  hyphaB. 

In  colonies  with  surface  strict  or  velvety  (consisting  of  conidiophores 
only)  there  is  a  succession  of  structures  from  the  center  to  the  periph- 
ery. In  the  center  arc  conidiophores  with  ripe  conidia,  marked  by  the 
colored  area.  This  shatles  into  a  wliite  margin  of  developing  conidio- 
phores and  coniiUal  fructifications,  wliile  the  extreme  margin  consists 
of  submerged  vegetative  hypha?.  The  relative  width  of  these  areas 
and  rate  of  the  development  of  conidiophores  and  colored  conidia  give 
characteristic  appearances  to  colonies  of  particular  species.  In  some 
there  is  a  broad  submerged  vegetative  border,  then  a  similar  white 
band  of  developing  conidiophores.  In  others  the  area  of  colored 
conidia  extends  so  closely  to  the  margin  that  the  white  border  is 
barely  discernible.  In  the  floccose  species  aerial  mycelium  often  ex- 
tends as  rapidly  at  the  margin  as  does  the  submerged  part.  In  such 
cases  the  area  of  coloration  follows  the  expansion  of  the  colony  more 
slowly. 

Although  close  resemblances  in  culture  are  not  uncommon,  the 
relative  development  of  these  areas  is  quite  typical  and  often  sharply 
distinctive  of  species.  Colonies  showing  a  broad  submerged  and  wliite 
margin  usually  spread  over  wide  areas  of  the  substratum,  whereas 
those  bearing  ripe  fruit  to  the  very  edge  of  the  growing  colony  rarely 
develop  beyond  restricted  areas. 

The  gross  characters  already  discussed  have  purely  specific  value, 
or  may  even  be  more  closely  restricted  as  characters  distinguishing 
particular  ecads  of  species.  The  generic  characters  and  very  impor- 
tant specific  characters  are  microscopic,  and  include  cell  relations  and 
details  of  spore  formation. 
8108— Bull.  118—10 2 


18  CULTURAL  STUDIES  OF   SPECIES  OF   PENICILLIUM. 

Conidiophore. — The  essential  data  as  to  conidiophores  are  their 
length,  septation,  the  diameter  of  their  cells,  and  especially  their  origin 
and  relation  to  the  substratum  and  to  each  other.  Although  ex- 
tremes of  variation  in  length  of  conidiophore  may  be  very  marked  in 
any  culture,  the  majority  of  conidiophores  in  any  such  culture  ap- 
proximate an  average  length.  This  length  to  be  most  reliable  must 
be  taken  from  the  origin  in  another  hypha  to  the  lowest  branch  of  the 
fructification.  If  the  conidial  fructification  were  counted  into  the 
length,  the  length  of  conidiophore  would  in  many  cases  be  doubled 
with  the  maturing  of  the  spores.  The  actual  length,  however,  is  little 
changed  with  such  maturity.  Valid  data  on  these  points  can  be 
secured  in  many  species  only  by  direct  observation  of  the  undis- 
turbed colony  in  the  air  under  the  microscope,  instead  of  by  the  study 
of  fluid  mounts.  This  is  equivalent  to  saying  that  Petri  dishes,  or 
other  vessels  which  can  be  uncovered  for  study,  must  be  used.  The 
student  must  expect,  therefore,  to  make  many  cultures  and  jeopardize 
the  purity  of  one  such  culture  every  time  he  undertakes  its  proper 
examination. 

Conidial fructijication. — For  lack  of  a  better  term  conidial  fructifica- 
tion may  be  defined  as  including  the  chains  of  conidia,  the  conidiiferous 
cells,  and  the  branches  bearing  them  back  to  their  junction  with  the 
conidiophore  proper.  In  a  species  of  PenidUium  fructifications  vary 
greatly  in  detail,  so  that  satisfactory  illustration  becomes  difficult. 
Comparison  of  large  numbers  of  fruiting  branches  in  many  cultures  of 
the  several  species  establishes  specific  types  of  appearance  which  can 
be  shown  in  a  conventionalized  series  of  sketches  for  each  species. 
The  data  found  of  value  have  been  the  mode  of  brandling,  the  meas- 
urements of  branches  and  conidiiferous  cells,  the  relation  of  these  to 
each  other,  the  arrangement  of  the  chains  of  conidia  with  reference  to 
each  other,  and  the  measurement  and  appearance  of  the  fructification 
as  a  whole.  These  penicillate  fructifications  vary  from  close  columns 
of  spores  arising  from  single  verticils  of  cells  borne  directly  upon  the 
apices  of  the  conidiophores  to  widely  spreading  "brooms"  whose 
numerous  divergent  chains  arise  from  verticillately  or  complexly 
arranged  branches  from  the  origin^il  conidiophores.  In  the  study  of 
these  conidial  fructifications  mounting  in  fluid  commonly  greatly  dis- 
arranges these  complex  masses  of  branches  and  spore  chains.  Direct 
observation  of  the  undisturbed  colony  is  therefore  essential  to  a  cor- 
rect conception  of  the  habits  of  the  species,  though  the  details  of 
branching  and  spore  bearing  must  be  sought  in  fluid  mounts. 

The  term  "  conidiiferous  cell "  is  used  for  the  cell  at  the  base  of  every 
chain  of  conidia  in  preference  to  "sterigma, "  as  often  used,  or  to 
"basidium"  as  used  by  Stoll."  The  term  was  proposed  by  Professor 
Atkinson  as  meeting  the  objection  that  basidium  implies  a  relation- 
ship net  justified  in  fact,  while  sterigma  usually  designates  not  a  spore- 


PHYSIOLOGICAL  EFFECTS  UPON   CULTURE   MEDIA.  19 

producing  cell  but  a  spore-bearing  process  of  a  cell,  hence  is  properly 
applicable  to  the  narrowed  apices  of  these  very  cells.  In  the  Latin 
descriptions  the  term  "basidium"  has  been  retained,  however. 

Conidia. — The  usual  data  with  reference  to  the  conidia  are  of  equal 
service  in  our  cultural  studies,  viz,  shape,  size,  color,  arrangement, 
markings,  mode  of  germination,  and  conditions  of  growth.  A  record 
of  the  changes  in  color  at  diflFereht  ages  of  the  colony  is  essential. 
Completeness  in  observation  is  as  necessary  in  spore  characters  as 
elsewhere.  The  variations  in  the  size  of  conidia  are  notable  in  some 
species;  in  others  conidia  appear  to  be  either  globose  or  elliptical,  even 
in  the  same  chain.  Where  the  outer  cell  wall  is  marked  or  spiny  these 
markings  often  do  not  appear  until  the  conidia  are  fully  mature. 

PHYSIOLOGICAL   EFFECTS    UPON    MEDIA. 

Certain  physiological  effects  of  fungous  growth  which  are  incident 
to  the  use  of  ordinary  culture  media  have  been  found  to  be  reliably 
characteristic  of  species.  Some  of  these  reactions  are  so  conspicuous 
as  to  aid  greatly  in  separating  nearly  related  organisms.  Such  re- 
actions as  have  been  found  uniform  and  definite  or  unique  are  intro- 
duced into  the  technical  diagnoses  of  species  in  this  paper.  Other 
physiological  data  are  appended  as  accessory  cultural  information. 

Among  the  data  observed  in  repeated  series  of  comparative  culture 
are  the  following:  Odor,  litmus  reaction  of  medium  at  different 
stages  of  growth,  liquefaction  of  gelatin  media,  the  production  of 
coloring  substances  in  the  media,  the  changes  produced  in  milk,  and 
the  production  of  drops  of  transpired  fluid  upon  surface  of  colony. 
Among  the  accessory  data,  observations  upon  carbon  assimilation, 
upon  proteolytic  reactions,  and  upon  the  production  of  enzymes  have 
been  made  for  certain  species. 

Odor. — The  production  of  definite  odors  by  colonies  is  confined  to  a 
small  number  of  species,  and  even  among  these  often  to  particular 
media.  When  definitely  present  it  is  a*  character  immediately  recog- 
nizable and  in  certain  species  diagnostic;  in  others  it  associates  the 
organism  at  once  with  a  particular  group  of  species. 

Litmus. — The  use  of  an. indicator  in  the  medium  gives  in  very 
many  species  a  sharp  reaction.  The  value  of  this  reaction  is  more  nar- 
rowly restricted,  however,  than  is  indicated  in  previous  papers 
(Thom  "  2«- ")  'pi^g  introduction  of  sterilized  litmus  or  azolitmin  into 
complex  media  brings  contradictory  results  when  the  composition  of 
the  medium  is  slightly  altered.  If,  for  example,  a  solution  of  pure  gela- 
tin in  distilled  water  be  used  as  a  nutrient,  the  reactions  are  definitely 
alkaline,  with  very  few  exceptions,  which  are  just  as  definitely  acid. 
The  gelatin  solution  itself  is  acid.  The  alkaline  reaction  indicates 
that  the  products  of  the  digestion  of  gelatin  in  such  cases  are  of  alka- 
line nature.     If,  however,  the  gelatin  solution  be  neutralized  and 


20  CULTURAL  STUDIES   OF   SPECIES  OF   PENICILLIUM. 

5  per  cent  cane  sugar  be  added,  an  equally  large  majority  of  the  same 
species  cause  an  acid  reaction.  Some  few  remain  alkaline.  These 
species  growing  upon  cane  sugar  produce  acids  in  quantity  both  to 
neutralize  the  alkaline  products  of  the  decomposition  of  gelatin  and 
to  change  the  reaction  of  the  mass.  The  litmus  reaction  has  been 
found  generally  reliable  in  a  medium  composed  of  15  per  cent  gelatin 
in  distilled  water,  in  lactose-peptone  gelatin  after  Conn's*  formula, 
and  in  potato  agar  (hiring  four  years  of  cultural  work  where  many 
successive  lots  of  media  Iiave  involved  the  use  of  materials  from 
different  sources.  In  complex  media  the  decomposition  products  of 
organic  nitrogenous  constituents  and  those  of  carbohydrates  tend 
either  to  neutralize  each  other  or  to  intensify  the  reactions,  but  add 
greatly  to  the  difficulties  of  analyzing  the  results.  The  litmus 
reaction  therefore  may  be  used  with  the  simplest  organic  media,  but 
is  best  applicable  to  cultures  in  synthetic  media  where  analysis  of  the 
results  is  feasible. 

Gelatin. — The  liquefaction  of  gelatin  media  by  an  organism  in  cul- 
ture shows  its  ability  to  produce  a  particular  form  of  proteolysis. 
This  reaction  is  so  conspicuous  and  so  adaptable  to  cultural  use  that 
it  has  been  recorded  in  all  cultural  studies.  Various  investigators 
have  pointed  out  the  limitations  of  this  reaction.  In  cultivation 
nearly  all  of  the  species  of  Penicillium  have  been  found  to  grow  well 
upon  a  15  per  cent  solution  of  gelatin  alone  in  distilled  water.  Com- 
parison of  the  results  in  this  medium  with  liquefaction  of  several  forms 
of  gelatin  media  experimented  with  showed  essential  agreement. 
Since  the  advantages  all  lie  in  the  simplification  of  formulae,  the  data 
as  to  liquefaction  by  these  species  have  been  compiled  from  series  of 
parallel  cultures  of  all  the  species  upon  15  per  cent  gelatin  in  dis- 
tilled water. 

The  value  of  this  reaction  is  measurably  vitiated  by  the  fact  that 
any  species  which  can  subsist  upon  gelatin  alone  must  be  capable  of 
more  or  less  proteolytic  action  upon  it.  The  results  of  observation 
confirm  this  statement.  The  value  of  these  observations  therefore 
depends  upon  the  indication  of  the  comparative  rate  of  activity  of 
different  species  in  inducing  proteolysis,  not  upon  the  presence  or 
entire  absence  of  this  action.  Numerous  tests,  involving  a  range  from 
15°  to  25°  C,  show  that  slight  changes  of  temperature  affect  this 
reaction  only  to  the  extent  to  which  they  affect  the  rate  of  growth, 
but  do  not  disturb  the  comparative  value  of  the  data  obtained. 
Within  this  range  of  temperature  the  most  active  species  produce 
liquefaction  in  5  to  8  days,  other  vigorous  liquefiers  require  8  to  15 
days,  while  many  species  producing  no  liquid  or  but  traces  of  soften- 
ing in  15  days  produce  a  gradual  liquefaction  in  the  succeeding  2  to  4 
weeks.  In  studying  species  of  PeniciUium,  liquefaction  of  gelatin  has 
been  found  to  separate  such  species  as  produce  this  proteolysis  within 


PHYSIOLOGICAL  EFFECTS  UPON   CULTURE   MEDIA.  21 

the  time  needed  for  other  observations  upon  these  cultures — about  15 
days.  Where  Hquefaction  occurs  (hiring  the  active  growth  of  the 
colony  it  definitely  indicates  the  secretion  of  ectoenzyms  capable  of 
this  digestion.  Long-deferred  liquefaction  may  result  from  such 
secretion  or  from  the  liberation  of  endoenzyms  by  the  disorganization 
of    mycelium. 

Color  'production. — Certain  species  cause  marked  changes  in  color 
in  some  of  the  substrata  in  which  they  grow.  Such  reactions  are 
usually  produced  upon  certain  media  and  not  on  others.  They  are 
therefore  selective  reactions.  For  example,  one  species  produces  a 
bright  yellow  color  in  media  containing  milk  sugar,  such  as  milk  or 
peptone-milk  sugar-gelatin;  but  no  color  in  plain  potato  agar.  Some 
of  these  colors  are  soluble  in  alcohol  to  form  brightly  colored  solu- 
tions. The  chemical  nature  of  these  substances  has  not  been  studied, 
but  the  reactions  themselves  have  been  thoroughly  tested  for  a  few 
species.  In  these  species  color  production  uniformly  follows  the 
proper  culture  conditions. 

Production  of  fluid  upon  surface  of  colony. — The  presence  upon  the 
surface  of  the  colony  of  lai^e  drops  of  transpiration  water  which  may 
be  colorless  or  brightly  colored  by  excreted  products  is  common. 
Although  a  transient  character  dependent  to  some  measure  upon 
the  humidity  of  the  air  in  the  culture  vessel  for  its  prominence,  there 
is  much  difference  between  species  in  this  particular. 

Technical  descriptions  of  these  fungi  have  always  been  based, 
theoretically,  upon  morphology  only.  In  such  descriptions  the 
host  or  substratum  has  been  more  or  less  definitely  indicated.  Where 
the  organisms  are  parasites  or  saprophytes  closely  restricted  to  par- 
ticular substrata  and  conditions,  this  practice  can  perhaps  be  justly 
supported.  With  omnivorous  cosmopolitan  saprophytes  cultural 
study  quickly  shows  morjihology  to  be  affected  greatly  by  changed 
conditions  or  altered  composition  of  media.  Further,  very  many 
of  these  organisms  have  no  known  special  habitat.  A  description 
based  upon  a  specimen  of  one  of  these  species  found  in  the  field, 
without  comparative  culture,  might  possibly  contain  some  pecuhar 
character  which  would  identify  a  specimen  of  the  same  species  when 
next  found,  but  this  is  only  a  possibility.  In  practice  the  descrip- 
tions we  have  are  nearly  useless. 

The  introduction  of  cultural  study  carries  with  it  the  necessity  of 
recognizing  at  least  the  most  striking  physiological  data.  With 
these  species  of  PenidUium  such  data  have  been  found  as  reliable  as 
morphology.  Further,  the  careful  definition  of  moiphology  and 
reactions  upon  specified  substrata  under  known  conditions  is  only 
amplifying  and  rendering  definite  the  two  items — habitat  and  local- 
ity— which  have  always  been  included  in  ])lant  descriptions.  In 
recent  years  the  students  of  ecology  and  of  experimental  evolution 


22  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

have  shown  these  to  be  much  more  important  in  flowering  plants 
than  was  formerly  supposed.  The  introduction  of  these  characters 
is  thus  in  full  conformity  with  the  best  systematic  practices  of 
to-day,  where  the  life  history  of  the  species  is  investigated  as  fully  as 
possible. 

CULTURE  MEDIA. 

In  developing  the  descriptions  presented  the  following  media  have 
been  used : 

1.  Potato  agar.  This  was  made  as  described  in  another  paper  *^ 
(p.  7).  Essentially  this  is  simply  a  potato  infusion  to  which  agar- 
agar  is  added  in  any  proportion  desired.  Since  this  medium  is  free 
from  sugar  and  very  dilute  in  nutrient  value,  sugar  may  be  added  to 
cultivate  certain  species. 

2.  Bean  agar.  The  directions  for  making  bean  decoction  were 
obtained  from  Maz6  at  the  Pasteur  Institute  in  Paris.  Common 
white  beans  are  heated  in  five  volmnes  of  water.  Boiling  is  stopped 
just  before  the  swelling  of  the  cotyledons  would  rupture  the  seed 
coats.  This  gives  a  clear,  slightly  yellowish  liquid  which  filters 
readily  yet  contains  sufficient  nutrients  to  grow  many  species  nor- 
mally. Agar  may  be  added  as  desired.  Since  this  decoction  is 
poor  in  available  carbon,  the  addition  of  sugar  is  often  desirable  for 
many  species. 

3.  Peptone-milk  sugar-gelatin,  as  described  by  Conn.*  This  stand- 
ard bacteriological  formula,  with  and  without  litmus,  was  used  in 
developing  the  original  draft  of  these  descriptions. 

4.  Fifteen  per  cent  gelatin  in  distilled  water.  As  a  stock  medium, 
this  was  used  without  neutralization.  Comparative  cultures  neu- 
tralized with  NaOH  seemed  less  adapted  for  most  species  than  un- 
changed gelatin.  With  few  exceptions  the  common  species  of 
PenidUium  grow  readily  in  plain  gelatin  and  give  the  same  reaction 
as  in  Conn's  more  complex  formula. 

5.  Synthetic  media.  A.  W.  Dox,  chemist  to  this  investigation, 
has  modified  Czapek's  formula  for  a  synthetic  medium,  intended  to 
present  in  a  nearly  neutral  solution  unaffected  by  sterilization  the 
elements  necessary  for  fungous  growth.  In  stock  form  neither  nitro- 
gen nor  carbon  is  presented  in  this  fluid.  It  has  consequently  been 
found  an  excellent  means  of  testing  the  availability  of  these  elements 
in  various  forms.  For  convenience  this  will  be  referred  to  as,Dox's 
solution,  the  formula  of  which  is — 

Distilled  water centimeters. .  3, 000.  0 

Magnesium  sulphate grams. .  1.  5 

Dipotaasium  phosphate  (K2HPO4) do 3.0 

Potassium  chlorid do 1.5 

Ferrous  sulphate do. ...  .03 


NOMENCLATURE.  Sd 

In  the  work  here  reported  nitrogen  was  added  as  sodium  nitrate,  6 
grams.  Parallel  experiments  in  which  monopotassium  phosphate 
(KH2POJ,  which  gives  a  perfectly  clear  solution  of  a  strongly  acid 
reaction,  was  substituted  for  dipotassium  phosphate,  KgHPO^,  gave 
no  advantages  in  culture  to  offset  the  advantages  of  a  neutral  medium 
except  the  disappearance  of  the  traces  of  precipitated  magnesium 
phosphate.  The  availability  of  carbon  in  any  organic  form  can  be 
tested  readily  in  this  solution.  Solidified  media  are  obtained  by  the 
addition  of  agar. 

NOMENCLATURE. 

THE    GENERIC    NAME. 

The  generic  name,  PenidUium  Link,  is  held  in  this  paper  in  its 
hyphomycete  sense  to  designate  all  species  which  continue  to  propa- 
gate themselves  for  an  indefinite  number  of  generations  by  penicil- 
late  asexual  fructifications.  Such  grouping  does  not  imply  the 
author's  belief  in  the  phylogenetic  relationship  of  all  such  forms. 
The  penicillate  type  of  fructification  is  a  definite  character  which 
binds  together  in  this  way  into  a  ''form-genus"  a  large  number  of  cos- 
mopolitan and  omnivorous  saprophytes,  very  few  of  which  are 
known  to  produce  sexual  fructifications.  Within  tliis  heterogeneous 
group,  several  series  of  forms  possessing  particular  groups  of  char- 
acters have  been  separated  and  generic  names  have  been  based  by 
some  workers  upon  such  segregation.  Wehmer^*  founded  the  genus 
Citromyces  upon  two  such  species  causing  citric  acid  fermentation  in 
sugar  media,  with  its  morphological  basis  in  the  presence  of  a  single 
whorl  of  conidia-bearing  cells  at  the  apex  of  the  conidiophore.  Fur- 
ther study  shows  that  the  presence  of  tliis  character  alone  would 
group  together  forms  not  so  closely  related  to  each  other  as  to  other 
species  of  Penicillium  lacking  this  character.  It  seems  best,  there- 
fore, to  use  the  name  Penicillium  to  designate  the  entire  group  and 
leave  further  investigation  to  establish  permanent  genera  when 
really  genetic  relationships  shall  have  been  discovered. 

NOMENCLATURE   OF   SPECIES. 

In  considering  the  actual  problem  of  nomenclature  of  species 
several  positions  may  be  taken.  -It  is  not  difficult  to  find  published 
descriptions  of  single  species  in  this  genus  which  are  sufficiently 
indefinite  to  include  a  large  percentage  of  all  the  known  species.  As  a 
rule,  the  morphological  characters  of  the  various  forms  under  culti- 
vation would  not  exclude  them  from  half  a  dozen  of  the  older 
'  species  so  far  as  current  descriptions  go.  In  many  cases  there  would 
not,  however,  be  the  least  reason  for  the  adoption  of  one  name  in 
preference  to  another.     Shall  the  investigator  adopt  for  his  material 


24  CULTURAL  STUDIES  OP  SPECIES  OF  PENICILLIUM. 

a  name  previously  used  when  he  has  little  or  no  reason  to  believe  that 
he  has  the  organism  originally  described  under  that  name  ?  Granting 
the  apparent  impossibiUty  of  contradiction,  he  might  find  it  a  safe 
practice.  If,  however,  contradiction  should  arise,  the  position 
becomes  entirely  indefensible.  In  case  there  is  a  fair  probability  that 
the  forms  are  identical,  the  use  of  the  old  name  may  be  justified  per- 
haps without  direct  proofs.  The  alternative  position  calls  for  the 
complete  description  of  the  form  studied  and  its  distribution  in  cul- 
ture to  different  centei-s  of  cultural  work  under  a  new  specific  name 
associated  with  this  definite  material  and  description.  It  can  not  be 
contended  that  these  organisms  are  new  to  science,  for  it  is  entirely 
possible  that  certain  of  them  have  formed  the  basis  for  already  pub- 
lished descriptions.  This  course  can  be  justified  by  the  contention 
that  such  publication  applies  to  definite  material,  available  for 
examination,  culture,  and  comparison  by  others,  and  that  there  is  less 
probability  of  confusion  from  such  pubhcation  than  would  ensue 
from  the  use  of  names  long  published,  without  any  real  evidence  of 
the  identity  of  the  organisms  with  those  originally  studied.  After 
careful  examination  of  all  material  available  and  consultation  with 
many  workers  in  closely  similar  fields,  new  names  are  attached  to 
such  forms  as  by  continued  cultural  study  appear  to  be  sharply 
marked  species  but  not  identifiable  by  older  descriptions. 

The  name  P.  glaucum  is  not  used.  Careful  examination  of  literature 
and  of  cultural  material  from  several  sources,  together  with  confer- 
ences and  correspondence  with  investigatoi-s  in  numerous  laboratories, 
does  not  afford  evidence  as  to  what  form  was  originally  used  by  Link 
or  even  secondarily  by  Brefeld  ^  under  this  name.  The  name  as  used 
at  present  seems  to  be  applied  collectively  to  the  common  green 
forms  which  under  examination  are  quickly  found  to  be  not  one  but 
several  species.  Further  study  may  give  some  indication  as  to 
where  the  name  really  belongs,  but  until  that  time  there  is  little 
profit  in  applying  it  to  any  particular  form.  It  might,  perhaps,  be 
withheld  until  some  worker  succeeds  in  repeating  Brefeld's  classic 
studies  in  ascus  production,  and  then  applied  to  the  form  so  found. 

The  present  paper  is  not  intended  to  be  a  monograph  of  the  genus. 
It  is  presented  as  a  report  covering  several  thousands  of  cultures 
of  a  group  of  common  forms,  in  the  hope  that  the  descriptions  and 
key  offered  may  be  useful  to  others.  The  author  has  included 
studies  of  as  many  authenticated  cultures  of  the  more  recently 
described  forms  as  it  was  possible  to  secure.  The  verified  cultures 
secured  have  already  been  fisted.  Much  assistance  and  advice  were 
freely  given  by  Dr.  C.  Wehmer.  On  the  other  hand,  it  was  impossi- 
ble to  secure  cultures  of  the  species  recently  described  by  Qude- 
mans,*'  or  those  listed  by  Dierckx;^  and  one  recently  described  by 
Peck'*  also  appears  to  be  lost.     No  claim  to  completeness  can  be 


THE  TYPE  SPECIES,  25 

made,  though  the  material  may  possibly  form  the  nucleus  of  a  real 
monograph  later. 

In  the  examination  of  the  early  literature  of  this  genus  the  author 
is  greatly  indebted  to  Dr.  C.  L.  Shear,  of  the  Bureau  of  Plant  Indus- 
try of  this  Department,  for  the  use  of  books  and  for  much  careful 
cooperation  in  examining  and  discussing  the  subject  of  types  and 
descriptions. 

THE   TYPE    SPECIES. 

Link^*  in  his  "Observationes,"  published  in  1809,  established  the 
genus  PeniciUium  to  include  the  common  green  molds  having  a 
penicillate  type  of  conidial  fructification — a  conidiophore  branching 
more  or  less  complexly  at  its  apex,  such  branches  becoming  or  being 
tipped  by  cells,  each  of  which  produces  a  chain  of  conidia.  The  whole 
produces  a  brush-like  appearance,  the  chains  of  conidia  serving  as  the 
hairs  or  bristles  of  the  brush.  This  generic  name  has  been  universally 
accepted  to  include  in  a  form  genus  all  species  reproducing  themselves 
indefinitely  by  such  conidial  fructifications.  However  doubtful  we 
may  be  as  to  the  forms  originally  examined,  there  is  no  question  that 
we  know  the  general  type  of  structure  which  Link  intended  in  his 
description  of  the  genus. 

Under  the  genus  PeniciUium,  Link  placed  three  species.  The  first 
species  listed  was  P.  glaucum,  but  the  description  given  is  equally 
applicable  to  many  different  forms.  This  was  noted  as  frequent  in 
decaying  bodies  and  said  to  be  most  closely  related  to  P.  expansum 
(the  third  species  listed),  of  which  he  suggests  his  material  may  have 
been  but  undeveloped  specimens. 

The  second  species,  P.  candidum,  is  described  as  producing  round 
colonies,  with  mycelium  and  spores  white,  upon  decaying  fungi  and 
herbs.  Although  many  authors  have  used  this  name  for  material 
from  different  sources,  and  Morini  ^^  has  described  an  ascigerous  form 
under  this  name,  there  has  been  no  means  of  determining  what  form 
Link  had  in  mind  when  writing  his  description.  Many  forms  will  pro- 
duce white  mycelium  and  spores  under  special  conditions,  while  but 
one  of  those  examined  has  been  shown  to  do  this  under  all  conditions, 
and  this  one  is  so  specialized  in  its  habit  as  to  be  excluded  by  Link's 
statement  of  habitat  from  the  application  of  this  name.  Numerous 
authors  have  suggested  that  the  P.  "candidu7n"  forms  are  probably 
colonies  of  species,  colorless  under  special  conditions,  but  green  under 
other  cultural  conditions. 

The  third  species,  P.  expansum,  is  slightly  better  described,  while 
its  habitat  is  primarily  given  as  rotten  fruit,  although  the  author  has 
manifestly  extended  his  use  of  the  name  to  forms  growing  upon  other 
substrata  which  he  believed  to  be  identical  with  the  fungi  grown  upon 
decaying  fruit. 


26  CULTURAL  STUDIES  OP  SPECIES  OP  PENIClLLTUM. 

On  page  19  of  his  "Observationes"  Link  describes  the  genus 
Coremium  with  a  single  species,  C.  glaucum,  whicli  he  specifies  as  f(»uncl 
upon  decaying  fmits.  This  fungus  can  be  traced  through  a  con- 
nected series  of  pubHcations  giving  descriptions  and  figures  which 
show  the  original  conception  to  include  (if  not  entirely  to  be  drawn 
from)  the  large  green  coremia  which  develop  upon  apples  and  related 
fruits  decaying  in  storage. 

This  organism  is  figured  by  Greville^  as  Floccaria  glauca;  it  is  cited 
cited  by  Fries  ^  as  a  variety  of  Penicillium  crustaceum,  from  which  he 
has  "seen  it  originate  upon  apples  in  the  autumn."  It  is  twice 
referred  to  by  Corda  in  Icones  Fungorum  (Vol.  II,  p.  17);  he  cites 
Coremium  glaucum,  C.  citrimum,  and  C.  candidum  as  synonyms; 
again  in  Prachtflora  (p.  54,  taf.  XXV,  figs.  3,  4,  17,  18,  19,  20,  and  21) 
under  the  name  of  Coremium  vulgare  he  manifestly  had  this  same  spe- 
cies, although  he  groups  it  with  figures  which  appear  to  be  different 
organisms. 

These  figures  and  descriptions  cited  are  definite  enough  to  show 
that  workers  contemporary  with  Link  applied  the  name  Coremium 
glaucum  Link  to  the  coremiform  rot  of  the  apple.  As  a  result  of 
observations  of  living  material  Fries  considered  this  only  a  form 
of  Penicillium  crustaceum,  which  he  made  to  include  P.  glaucum  and 
P.  expansum  Link. 

The  present  writer  has  collected  this  fungus  upon  decaying  apples 
and  related  fruits  repeatedly  in  America;  also  upon  pears  and 
mespilus  in  Hanover^  Germany. 

Repeated  cultures  have  shown  that  the  ability  to  produce  coremia 
is  a  definite  character  of  this  species,  recognizable  under  many  con- 
ditions of  culture,  but  not  shown  under  other  conditions.  The  same 
culture  will  commonly  show  both  simple  penicillate  fructifications 
and  coremium  production.  The  species  must  therefore  be  regarded 
as  one  of  the  several  species  of  Penicillium  which  always  produce 
coremia  under  proper  cultural  conditions. 

Examining  Link's  species  of  Penicillium,  we  find  that  he  specifies 
P.  expansum  as  primarily  found  upon  rotten  fruit.  P.  expansum 
Link  clearly  included  Coremium  glaucum  Link,  therefore,  probably 
with  othei-s;  but  from  its  known  abundance  in  Germany  there  can 
be  little  question  as  to  this  organism  forming  in  part,  at  least.  Link's 
original  conception  of  this  species.  Later  (1824)  in  Species  Plan- 
tarum,  Tomus  VI,  page  70,  Link"  redescribes  P.  glaucum  and  includes 
in  it  the  P.  expansum  of  his  Observationes.  In  this  discussion  Link 
broadens  his  description  of  P.  glaucum  to  include  all  green  forms 
found  in  decaying  substances,  upon  the  assumption  that  all  such 
forms  are  but  a  single  species.  It  is  evident  that  the  earlier  descrip- 
tion P.  expansum  Link  included  this  species  with  sufficient  restriction 


PENTCILLIUM  EXPANSUM.  27 

to  justify  reviving  the  name  P.  expansum  Link  and  limiting  it  to  the 
peniciUium  rot  of  apples  alone. 

P.  expansum  Link  (in  part),  the  peniciUium  rot  of  apples,  would 
therefore  stand  as  the  type  species  of  the  genus. 

The  technical  rules  for  the  establishment  of  type  species  are  in  this 
way  satisfied.  The  kind  of  plant  used  by  Link  is  perfectly  well 
known.  To  say  just  which  forms  he  had  in  hand  may  be  impossible, 
but  the  form  we  are  discussing  was  certainly  one  of  them. 

EXPLANATION  OF  DRAWINGS. 

The  drawings  of  the  species  described  have  been  made  with  the 
Bausch  &  Lomb  camera  lucida  in  all  cases  except  such  as  are  marked 
diagrammatic  or  partly  diagrammatic. 

For  those  with  the  magnification  of  140,  the  lenses  used  were  the 
Bausch  &  Lomb  1-inch  ocular  and  the  two-thirds  objective;  for  those 
of  900  the  lenses  were  the  Bausch  &  Lomb  1-inch  ocular  and  the  one- 
eighth  objective;  for  those  of  1,400  the  lenses  were  the  Bausch  & 
Lomb  1-inch  ocular  and  the  one-twelfth  objective;  for  those  of  1,600 
the  Spencer  No.  12  compensating  ocular  and  the  Zeiss  3  mm.  apert. 
1.30  apochromatic.  This  apochromatic  was  also  used  in  a  few  cases 
at  a  magnification  of  900 ;  these  are  indicated  in  the  legends.  All 
figures  at  magnification  of  140  were  drawn  from  the  exposed  surface 
of  the  undisturbed  colony  in  Petri-dish  cultures.  All  other  figures 
were  made  from  fluid  mounts  or  hanging-drop  cultures  (for  spore 
germinations) . 

The  series  of  sketches  at  140  magnification  are  made  with  uniform 
methods,  so  that  comparison  of  species  in  culture  can  be  much  easier 
than  from  figures  made  at  different  magnifications. 

PENICILLIUM  EXPANSUM  Link. 

P.  expansum  Link  (in  part),  emended  Thom=penicillium  rot  of  apples  and  allied  fruits. 

Syn.  Coremium  glaucum  Link,  Observations,  p,  19;  Icones,  V,  fig.  31,  1809. 
Floccaria  glauca  Greville,  Scottish  Flora,  pi.  301,  figs.  1^. 
Penidllium  glaticum  Link  (in  part),  Species  Plantarum,  VI  (1824),  p.  70. 
Coremium  vulgare  Corda  (in  part),  Prachtflora,  p.  54,  PI.  XXV,  especially 

figs.  3,  4,  17,  18,  19,  20,  and  21. 
Possibly  P.  elongatum  Dierckx. 

Colonies  upon  gelatin  and  potato  or  bean  agar,  green  becoming  gray-green  and 
slowly  brown  in  several  weeks  (especially  when  exposed  to  light),  floccose,  with  concen- 
tric zones  tufted  with  short,  loose,  coremium-like  aggregations  of  conidiophores,  not 
over  1-2  mm.  in  height  except  in  old  cultures  containing  sugar,  broadly  spreading  with 
broad  white  margin  in  growing  colonies.  Reverse  somewhat  brown.  Conidiophores 
either  very  short  lateral  branches  of  aerial  hyphae  or  very  long  (1  mm.  or  more),  aris- 
ing singly  or  grouped  with  others  to  form  coremia.  Conidial  fructifications  consist 
of  1  to  3  main  branches  bearing  verticils  of  branchlets  supporting  crowded  whorla 
of  conidiiferous  cells,  130-200  by  50-60/x  at  base  in  cultures  without  sugar,  with  sugar 


28 


CULTURAL  STUDIES  OP  SPECIES  OP  PENICILLIUM. 


continuing  for  some  weeks  to  produce  great  numbers  of  conidia  which  come  to  form 
masses  perhaps  1  mm.  in  thickness.  Conidiiferous  cells  8-10  by  2-3/i.  Conidia 
elliptical  to  globose  2  by  3.3/t  or  3  3.4/(;  green,  homogeneous,  persisting  in  chains 
when  mounted.  Colonics  begin  to  liquefy  gelatin  slowly  after  about  10  days  and 
continue  until  it  is  completely  liquefied.  Grows  readily  and  rapidly  upon  all  com- 
monmedia. 

Occurs  characteristically  upon  decaying  apples  and  other  pomaceous  fruits,  where 
old  colonies  often  produce  coremia  1  cm.  or  more  in  length  and  very  lai^e. 

Collected  at  Ithaca  and  Geneva  (Eustace),  N.  Y.,  at  Middletown  and  Storrs,  Conn., 
upon  apples;  upon  pears  and  Mespilus  at  Hanover,  Germany.  Often  appears  as  a 
contamination  in  fungous  cultures. 


'id 


Fig.  l.—  Penicillium  ezpansum  Link:  a,  b,f,  branching  and  arrangoment  of  branches  of  conidial  fructifi- 
cation (X  900);  c,  d,  e,  conidiiferous  cells  and  conidial  chains  (X  900);  g,  h,  j,  k,  I,  sketches  of  fnictiflca- 
tion  (X  140);  m,  n,  o,  germination  of  conidia  (X  900);  r,  a,  sketches  from  photomicrographs,  showing 
in  »  loose  aggregations  of  conidiophores  Ijeginning  to  develop  in  zones  which  become  coremia  like  r 
(coremium  r  was  1  mm.  in  height;  X  35). 

Many  references  in  the  literature  to  P.  glaucum  Link  and  P.  crusta- 
ceum  (L.)  Fries  refer  to  this  species.  Unless,  however,  such  citations 
directly  refer  to  the  presence  of  coremia,  or  to  the  association  of  the 
organism  with  the  decay  of  pomaceous  fruits,  or  both,  there  is  no 
means  of  fixing  the  application  of  such  names  to  this  species.  Since 
Link"  in  1824  lumped  into  his  species  P.  glaucum  every  kind  of  green 


PENICILLIUM   ITALICUM.  29 

penicillium  for  which  he  could  find  references,  the  myth  ofe-^"the 
common  green  mohl"  seems  to  have  had  pretty  general  acceptance, 
although  we  find  here  and  there  a  protest  against  this  view. 

A  culture  of  this  species  can  always  be  obtained  from  apples  decay- 
ing in  storage,  upon  which  usually  well-developed  coremia  can  be 
found  if  proper  search  is  made.  The  wide  distribution  of  the  organism 
as  noted  above  and  as  seen  in  the  literature  justifies  belief  in  its 
general  distribution.  Once  carefully  observed  in  cultures,  the  in- 
vestigator will  usually  recognize  the  organism  on  sight  when  it  ap- 
pears in  his  cultures  even  as  a  contamination  of  other  species.  The 
odor  of  this  species  is  so  distinctive  as  to  assist  greatly  in  identifying 
it.  in  accidental  cultures.  It  does  not  produce  the  yellow  color  in 
the  substratum  as  described  by  Lindau  *"  for  P.  glaucum,  nor  are  its 
spores  globose  from  the  first  as  Wehmer^"  records  them  for  that 
species.  It  seems  so  very  well  characterized  as  to  justify  sharply 
separating  it  from  the  other  green  forms. 

CULTURAL  DATA. 

Color,  green  or  gray -green;  color  of  reverse,  yellowish  to  somewhat  brown;  color 
in  media,  colorless  or  yellowish  brown  (milk). 

Odor,  "fruity"  in  all  or  nearly  all  cultures. 

Fifteen  per  cent  gelatin  in  water,  good  growth,  with  loose  and  ill-defined  but  abun- 
dant coremia;  liquefaction  slow,  more  or  less  liquid  in  2  weeks;  litmus  reaction  alka- 
line or  neutral.  Potato  agar  and  bean  agar,  characteristic  colonies,  gray-green  with 
more  or  less  coremiform  bundles  of  conidiophores.  Potato  plugs,  characteristic 
colony. 

Raulin's  fluid  agar,  characteristic  colony  with  concentric  rings  of  broad  coremia. 
Raulin's  fluid,  characteristic  colony.  Cohn's  solution,  slight  growth,  few  coremia, 
brownish  below. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  coloring  up  to  50 
per  cent  sugar,  acid  reaction,  conidiophores  in  dense  areas  with  few  definite  coremia. 
Lactose  3  per  cent,  slow  development,  about  half  normal.  Levulose  3  per  cent, 
typical,  alkaline,  or  neutral  reaction.  Galactose  3  per  cent,  typical,  acid  reaction. 
Glycerin  3  per  cent,  slow  development,  increased  greatly  by  adding  sugar.  Potato 
starch  3  per  cent,  fair  growth,  no  coremia.     Butterfat,  rich  typical  growth. 

Milk,  typical  colonies,  coremia  in  a  ring  at  glass;  curdling  (0.25  per  cent  calcium 
chlorid  added)  in  1  week;  digestion  rather  slow;  color  in  milk  yellowish  brown. 

At  37°  C,  no  growth,  culture  grew  when  cooled;  at  20°  C,  good  growth. 

The  coremia  of  this  species  are  especially  characteristic  of  old  cultures  in  which 
drying  has  begun.  In  fluid  cultures  they  are  commonly  attached  to  the  glass  above 
or  at  the  very  top  of  the  fluid. 

PENICILLIUM  ITALICUM  Wehmer. 

Beitr.  z.  Kennt.  einh.  Pilze,  Jena,  1895,  p.  68,  t.  II. 

Colonies  on  plain  gelatin  and  potato  or  bean  agar  bluish  green,  becoming  gray -green 
when  old  (bright  bluish  green  on  cane-sugar  media  and  citrus  fruits),  broadly  spreading, 
aerial  portion  composed  at  the  broad  margin  almost  entirely  of  conidiophores,  but  be- 
coming slightly  floccose  in  the  center.  Reverse  of  colonies  dark  brownish,  often  almost 
black  in  media  containing  sugar.  Conidiophores  from  short  (100/t)  to  very  long  (GOOfi), 
averaging  perhaps  250/t,  arising  either  directly  from  substratum  or  as  branches  of 
aerial  hyphse.    Conidial  fructifications  up  to  300/z  or  more  in  length,  consisting  usually 


30 


CULTUEAL,  STUDIES  OF  SPECIES  OF  PENICILLIUM. 


of  a  main  branch  and  one  lateral  branch,  each  producing  a  whorl  of  branchlets  bearing 
crowded  verticils  of  conidiiferous  cells,  12-14  by  3/t.  Conidia  breaking  off  in  masses  in 
handling  old  cultures,  which  rise  in  clouds  when  shaken.  Pronounced  odor  in  cul- 
tures containing  cane  sugar.  Chains  of  conidia  loosely  divergent,  long;  conidia 
2-3  by  3-5/«,  cylindrical  to  elliptical  or  slightly  ovate,  clear  green  by  transmitted 
light,  very  variable  in  size  but  usually  within  the  limits  giVen.  Masses  of  spores  con- 
tinue to  increase  from  2  to  3  weeks.  Petri-dish  colonies  partially  and  slowly  liquefy 
gelatin  (12  to  20  days).     Numerous  white  sclerotia  are  produced  upon  the  surface  of 


Fig.  2. — PcnkfOium  italicum  Wehmer:  o,  6,  c,  d,  e,  /,  g,  types  of  branching,  formation  of  verticils  of 
conidiiferous  cells  and  conidial  chains  (a,  c,  f,  X  900;  6,  c,  d,  g,  X  1,400);  j,  k,  sketches  of  conidial  fructi- 
fications (X  140);  I,  m,  n,  swelling  and  germination  of  conidia  (X  900). 

the  medium  after  2  to  3  weeks'  growth,  especially  upon  fruits  and  other  acid  media 
rich  in  sugar. 

Cosmopolitan;  characteristic  of  decaying  oranges,  which  become  bright  blue-green 
with  this  mold  as  contrasted  with  the  olive-green  species  which  is  often  aasociate4 
with  it  upon  the  same  fruit.    These  contrasting  colors  are  illustrated  by  Wehmer.^' 


PENICILLIUM  DIGITATUM.  31 

In  his  discussion  of  this  fungus  Wehmer"  shows  that  the  blue-green 
rot  of  citrus  fruits  is  a  different  species  from  the  similarly  colored 
apple  rot.  His  description  seems  to  be  the  first  recognition  of  this 
species  as  different  from  the  green  molds  occurring  constantly  upon 
all  kinds  of  food.  This  species  has  been  discussed  as  P.glaucum  in 
recent  papers  (R.  E.  Smith/^  PowelP")^  where  its  agency  in  the  decay 
of  citrus  fruits  is  very  fully  considered. 

Cultures  were  obtained  by  the  writer  from  oranges  in  Hanover, 
Germany,  and  identified  by  the  describer.  Dr.  C.  Wehmer.  It  has 
since  been  repeatedly  observed  and  collected  in  America.  Pure  cul- 
tures can  always  be  secured  by  finding  decaying  oranges  in  the  mar- 
ket which  have  the  blue-green  areas  of  rot  just  beginning  to  appear 
upon  them.  These  areas  are  usually  blue-green  in  center  surrounded 
by  white  areas  which  are  usually  grouped  into  little  white  patches 
toward  the  vegetative  margin  and  the  whole  superficial  colony  sur- 
rounded by  an  area  of  soft,  watery  rot.  Very  often  such  colonies 
when  older  become  much  contaminated  with  the  olive-colored  rot, 
given  in  this  paper  as  P.  digitatum. 

CULTURAL  DATA. 

Color,  clear  bluish-green  on  sugar  media,  shades  of  gray-green  without  sugar;  reverse 
of  colony  commonly  brownish  in  areas;  color  in  media  none  or  slight. 

Odor  distinct  upon  media  containing  cane  sugar,  none  on  lactose  or  media  free  from 
sugar. 

Fifteen  per  cent  gelatin  in  water,  medium  growth;  liquefaction,  none  until  several 
weeks  old,  then  partial  in  acidified  cultures;  litmus  reaction  fairly  alkaline.  Potato 
agar  with  lactose,  rather  thin  gray-green  colonies,  not  vigorous,  acid  reaction.  Potato 
plugs,  pale  but  characteristic.  Raulin's  fluid,  typical  colonies.  Cohn's  solution, 
germination  only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  colonies  up  to  50  per 
cent  with  alkaline  reaction.  Lactose  3  per  cent,  slow  growth  but  typical,  with  acid 
reaction.  Lactic  acid  0.9  per  cent,  small  growth.  Levulose  3  per  cent,  not  normal. 
Galactose  3  per  cent,  medium  growth,  acid  reaction.  Glycerin  3  per  cent,  no  growth; 
growth  began  when  sugar  was  added.     Butterfat,  growth  slow. 

Milk,  good  growth;  curdling  (0.25  per  cent  calcium  chlorid  added),  ten  days;  diges- 
tion partial  and  slow;  color  slowly  brownish  or  yellow-brown. 

At  37°  C . ,  killed ;  at  20°  C . ,  good  growth . 

PENICILLIUM  DIGITATUM  Saccardo. 

The  olive-green  orange  rot,  P.  digitatum  Sacc,  in  Mycotheca  Italica,  No.  986,  Her- 
barium U.  S.  Department  of  Agriculture;  in  Sylloge  Fungorum,  Vol.  IV,  p.  79;  in 
Fungi  Italici,  No.  894. 

=P.  olivaceum  Wehmer,  Beitr.  z.  Kennt.  einheim.  Pilze,  pp.  73,  t.  II,  Jena,  1895. 
?MiLcor  casspitosus  L.,  in  Species  Plantarum  (1753),  II,  p.  1186,  based  upon 

Micheli,  tab.  91,  fig.  3. 
?Monilia  digitata  Fries,  Systema  Mycqiogicum,  III,  p.  411. 

Colonies  on  sugar  gelatin  and  potato  or  bean  agar  grayish  olive,  irregularly  shaped 
from  the  unequal  growth  and  branching  of  rather  few  hyphae,  aerial  portion  consisting 
only  of  very  short  conidiophores  and  conidia.  Reverse  of  colony  commonly  shows 
brown  to  black  colors.    Conidiophores  rising  directly  from  the  substratum.  30-100  by 


32  CULTURAL  STUDIES  OF   SPECIES  OF   PENICILLIUM. 

4-bfi,  usually  very  Hhort.  ,Conidial  fructification  a  few  tangled  conidial  chains  up  to 
160/t  in  length,  borne  upon  conidiiferous  cells  13-16  by  S-A/i.  Conidia  cylindrical  to 
almost  globose,  4-7  by  6-8/i  (at  times  6  by  10/t),  often  uneven  in  size  and  shape  in  the 
same  chain.  Colonies  do  not  liquefy  sugar  gelatin  except  at  times  partially  in  cultures 
three  weeks  old  or  more.  Litmus  reaction  acid.  Grows  readily  on  organic  media,  but 
shows  a  very  pronounced  aflinity  for  such  media  with  high  percentages  of  sugar,  in 
which  it  produces  a  strong  odor.  Refused  to  grow  in  synthetic  media  containing  nitro- 
gen as  sodium  nitrate. 

Cosmopolitan  uj)on  citrus  fruits,  distinguished  from  P.  italicum  by  the  sharp  con- 
trast of  its  olive  color  with  the  blue  of  the  other.  Collected  in  Hanover  and  verified 
by  Dr.  C.  Wehmer.  Received  from  Prof.  P.  H.  Rolfs  in  I'lorida.  Seen  upon  decaying 
oranges  everywhere.  Pure  cultures  can  always  be  secured  from  the  common  market 
fruits. 

Nomenclature, — Wehmer^*  (1895)  gives  the  first  adequate  discus- 
sion of  the  decay  of  citrus  fruits  by  the  agency  of  species  of  Penicil- 
lium  in  which  the  forms  found  were  shown  to  be  distinct  species 
associated  constantly  with  these  fruits  instead  of  common  green 
species  accidentally  occurring  upon  these  fruits.  For  the  olive- 
green  form  the  name  proposed,  P.  oHvaceum,  is  descriptive,  but  had 
already  been  used  by  Corda  (Icones,  III,  p.  12,  t.  II,  fig.  35)  for 
a  species  afterwards  transferred  to  Ilormodendrum.  The  name 
P.  olivaceum  Wehmer  is  therefore  not  tenable  by  present  rules  of 
nomenclature, 

P.  digitatum  is  the  name  used  by  Saccardo,  as  is  shown  by  the 
specimens  distributed  by  him  (Mycotheca  Italica,  No.  986)  previous 
to  Wehmer's  work.  In  his  description  and  synonymy  he  cites  the 
name  from  Fries  (Systema  Mycologicum,  p.  411),  where  it  appears  as 
Monilia  digitata,  transferred  by  Saccardo  to  Penicillium.  But  F'ries 
cites  his  use  of  the  name  from  Persoon  '"  (Synopsis  Fungorum,  p.  693), 
who  bases  his  description  of  Monilia  digitata  upon  Micheli's  ^*  figure 
and  description  (Nova  Plantarum  Genera,  p.  213,  pi.  91,  fig.  3). 
Here,  under  the  name  of  Aspergillus,  Micheli  figures  and  describes  as 
No.  8  a  fungus  which  may  be  taken  for  a  Penicillium,  which  is  said  to 
have  been  found  upon  semi-putrid  lemons.  This  figure  is  cited  by 
Linnaeus  in  Species  Plantarum  (1753),  II,  p.  1656,  as  the  basis  of  his 
Mucor  csespitosus. 

If  we  accept  Saccardo's  citations  as  correctly  giving  the  origin  of 
the  name  and  tracing  it  back  to  Micheli,  we  must  abandon  the  name 
he  uses  and  adopt  the  name  given  by  T^innaeus — Mucor  caesjntosus  = 
P.  csespitosum  (L.).  Careful  scrutiny  of  Micheli's  figure  gives  no 
possible  means  of  identification.  Aside  from  identificatiop  from  pre- 
served specimens  which  are  not  cited  by  the  authors  in  any  case, 
there  is  little  possibility  of  showing  what  any  of  the  authors  com- 
monly cited,  for  this  species  mayjiave  had  until  we  come  to  the 
description  given  by  Saccardo  in  the  Sylloge  (FV,  p.  79),  and  the 
material  distributed  by  him  in  Mycotheca  Italica,  which  I  have  seen 
and  which  is  certainly  this  fungus.  I  have  therefore  continued  the 
use  of  Saccardo's  name  in  this  case  because  he  has  given  the  first 


PENICILLIUM  DIGITATUM. 


33 


description  based  upon  tangible  material,  but  have  rejected  his 
citations.  The  following  are  the  principal  citations  which  have  been 
given  as  referring  to  this  species: 

Micheli,  Genera  Plantarum  (1729),  pi.  91,  fig.  3. 

Linnaeus,  Species  Plantarum  (1753),  II,  p.  1656. 

Persoon,  Observations,  p.  41. 

Persoon,  Synopsis  Fungorum,  p.  693,  Monilia  digitata. 

Fries,  Systema  Mycologicum  (1829),  III,  p.  411,  Monilia  digitata. 

Saccardo,  Fungi  Italici  (1881),  No.  894,  and  Syllogc,  IV,  p.  79,  P.  digitatum. 

Wehmer,  beitr.  z.  Kennt.  einheim.  Pilze,  p.  73,  taf.  II. 


Fig.  3. — Pemcillium  digitatum  Saccardo:  a,  whole  conidiophore  and  fructification;  6,  c,  d,  e,  types  of  branch- 
ing and  formation  of  conidia;  m,  n,o,  germination  of  conidia;  all  X  900. 

The  only  reason  for  retaining  the  name  P.  digitatum  and  ascribing 
its  authorship  to  Saccardo  is  that  the  description  cited  is  the  first  one 
which  we  can  ascribe  definitely  to  material  of  this  species.  If,  as 
said  above,  his  citation  of  literature  proves  to  be  correct,  the  name 
he  gives  becomes  untenable  by  present  rules  of  nomenclature,  and 
must  be  changed  to  P.  csesjntosum  as  given  by  Linnaeus,  although 
there  is  no  evidence  that  Linnaeus  actually  examined  material  of  this 
species.  Since  there  is  more  or  less  doubt  about  usages  previous  to 
Saccardo,  and  since  the  name  given  by  Saccardo  has  become  asso- 
8108— Bull.  118—10 3 


84  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

ciated  with  studies  of  this  species  in  recent  hterature  (Powell,'" 
Smith  '*),  this  name  is  allowed  to  stand,  at  least  until  we  have  more 
information. 

CULTURAL  DATA. 

Color,  shades  of  olive  green;  reverse  brown  or  dark  brown,  especially  in  sugar  media; 
color  in  media,  none  or  slightly  yellowish. 

Odor,  associated  with  smell  of  rotting  oranges,  strongest  upon  cane-sugar  media. 

Fifteen  per  cent  gelatin  in  water,  weak  growth,  not  adapted  for  this  species;  lique- 
faction none,  slowly  accomplished  in  gelatin  to  which  sugar  is  added;  litmus  reac- 
tion acid.  Potato  agar  and  bean  agar,  colonies  as  described  above,  rather  poor  growth 
which  becomes  enormously  increased  upon  the  addition  of  cane  sugar.  Potato  plugs, 
excellent  growth.  Raulin's  fluid,  colony  colorless.  Cohn's  solution,  germination 
only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  germinated  only.  Lac- 
tose 3  per  cent,  germinated  but  no  growth.  Lactic  acid,  0.9  per  cent,  germinated 
only.  Levulose  3  per  cent,  germination  only.  Galactose  3  per  cent,  slight  growth. 
Glycerin  3  per  cent,  germination.  Potato  starch  3  per  cent,  slight  or  no  growth.  But- 
terfat,  germination  only.  Conidia  inoculated  into  this  fluid  with  various  sources  of 
carbon  germinated  but  never  developed  normal  colonies. 

Milk. — This  species  never  produced  vigorous  growth  upon  milk. 

At  37°  C,  killed;  check  at  20°  C.  grew  well. 

PENICILLIUM  ROQUEFORTI  Thom. 

U.  S.  Department  of  Agriculture,  Biu-eau  of  Animal  Industry,  Bui.  82,  pp.  35-36, 

fig.  2,  1906. 
Report  Storrs  Agricultural  Experiment  Station,  1905,  p.  111. 

Syn.  P.  glaucum,  various  authors— not  Link  or  Brefeld. 
Colonies  on  potato  agar  or  lactose  gelatin  quickly  turning  green,  becoming  a  dirty 
brown  when  old,  velvety  strict,  indeterminately  spreading  by  large  main  radiating, 
branching  hyphse,  giving  a  somewhat  uneven  or  indefinite  margin,  which  gets  a  white, 
fibrous,  almost  spider-web  appearance  from  its  alternation  of  submerged  parts  of 
hyphae  with  short  prostrate  aerial  loops.  Reverse  of  colony  yellowish  white. 
Conidiophores  arising  separately  and  in  acropetal  succession  from  the  growing  parts 
of  submei^ed  hyphae  (comparatively  few  aerial  parts,  but  some),  200-300/t  septate. 
Fructitication  90-120,«  or  at  times  160  by  30-60/t  at  broadest  place,  usually  appear- 
ing double  by  the  divergence  of  the  lowest  branch;  branchlets  ("basidiophores") 
irregularly  verticillate,  bearing  crowded  verticils  of  appressed  conidiiferous  cells 
(basidia),  9-11  by  2.5/i  with  long,  divergent  chains  of  conidia.  Conidia  bluish  green, 
cylindrical  to  globose,  smooth,  rather  firm  walled,  4-5/t  in  diameter,  germinating  by 
a  straight  tube.  Colonies  do  not  liquefy  sugar-gelatin,  though  they  soften  it  some- 
what. Fungus  on  plain  gelatin  or  potato  agar  changes  litmus  from  red  to  blue  very 
rapidly  and  strongly  almost  from  the  beginning  of  the  growth.  Fruiting  period  short, 
but  one  crop  of  spores  upon  the  mycelium.  Cosmopolitan  and  omnivorous,  or  nearly 
so.    Characteristic  of  Roquefort  and  related  types  of  cheese. 

The  mold  of  Roquefort  and  related  types  of  cheese  has  been  com- 
monly <lesignated  in  dairy  literature  as  P.  glaucum  Link.  Such 
citations  refer  to  this  fungus,  though  many  times  referring  to  it  as 
"the  common  green  mold."  Although  it  is  not  restricted  in  its 
habitat  to  cheese,  this  species  is  so  identified  with  the  ripening  process 
of  Roquefort  cheese  (in  which  pure  cultures  are  used)  that  any  one 


PENICILLIUM  ROQUEFORT!. 


35 


desiring  a  culture  of  this  species  can  always  obtain  it  by  purchasing 
cheese  of  this  type.  It  also  occurs  frequently  in  ensilage  and  is 
often  found  as  a  contamination  in  laboratory  cultures  of  other  fungi. 
The  species  is  well  marked  ])y  cultural  characters  from  which  it  can  be 
readily  recognized  when  once  studied.  Its  agency  in  the  ripening 
of  Roquefort,  Gorgonzola,  and  Stilton  cheeses  is  discussed  in  another 
paper  (Thom^^) 


CULTURAL   DATA. 


Color,  gray-green  to  clear  green,  becoming  brownish  when  old  if  exposed;  reverse 
colorless  or  cream;  color  in  media,  none,  or  in  some  cases  a  slight  and  evanescent 
greenish  tinge. « 

Odor,  none  (unless  accountable  for  some  odor  in  Roquefort  cheese). 


Fig.  4.—Penicillium  roqueforti  Thorn:  a,  part  of  conidiophore  and  of  base  of  fructification,  highly  mag- 
nified, showing  production  of  basidia  on  sides  as  weli  as  at  ape.x  of  basidiophore;  6,  c,  other  types  of 
branching;  d,  young  conidiophore  just  branching;  e,  f,  basidia  and  the  formation  of  conidia,  highly 
magnified;  g,  h,j,  diagrams  of  types  of  fructification  as  seen  under  low  power  (X  80);  fc,  I,  m,  n,  germina- 
tion of  conidia  and  new  conidia  produced  directly  on  the  first  h3rph£e.  (From  Bulletin  82,  Bureau  of 
Animal  Industry.) 

Fifteen  per  cent  gelatin  in  water,  good  growth,  not  heavy;  liquefaction,  none,  or 
partial  after  2  to  3  weeks  in  acidified  cultures;  litmus  reaction,  alkaline.  Potato  agar 
and  bean  agar,  gray-green,  loose,  becoming  dense  and  deep  green  when  sugar  is  added. 
Potato  plugs,  characteristic.  Raulin's  fluid,  very  dense,  deep  green  colonies.  Cohn's 
solution,  slight  growth. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  slow  growing  but  typical 
colonies  up  to  30  per  cent,  persistently  green,  not  acid.     Lactose  3  per  cent,  weak 

a  Weidemann'*  records  the  production  of  a  red  color  in  cane-sugar  solution  by  this 
species.     The  writer  has  not  obtained  this  color  in  his  cultures. 


36  CULTURAL  STUDIES  OP   SPECIES  OF  PENICILLIUM. 

growth,  evidently  carbon  deficiency.  Lactic  acid  0.9  per  cent,  better  than  lactose, 
not  typical  colony,  good  color.  Levulose  3  per  cent,  weak  colonies.  Galactose  3  per 
cent,  good  growth,  alkaline.  Glycerin  3  per  cent,  weak  growth.  Alcohol,  good 
growth,  becoming  brown  in  age.  Tartaric  acid,  very  slight  growth.  Butterfat,  rich 
growth. 

Milk,  growth  good,  alkaline  to  litmus;  curdling  (0.25  per  cent  of  calcium  chlorid 
added)  in  10  days;  digestion,  fairly  rapid;  color  in  milk,  none. 

At  37°  C,  no  growth;  grew  when  cooled;  check  at  20°  C,  good  colony. 

PENICILLIUM  PURPUROGENUM  O.  StoU. 

Beitr.  z.  morph.  u.  biol.  Char.  Penicillium,  Wurzburg,  1904,  p.  32,  t.  I,  fig.  6;  t.  Ill, 
fig.  2;  t.  IV,  fig.  3. 

Colonies  on  lactose  gelatin  and  potato  or  bean  agar,  gray-green  to  brown  or  olive, 
deeper  green  upon  cane-sugar  media,  closely  floccose,  almost  velvety  in  surface 
appearance,  spreading  slowly  over  the  substratum  and  producing  in  the  whole  mass 
of  medium  a  red  color.  In  acid  media  rich  in  sugar  secondary  floccose  mycelium  arises 
white  or  with  hyphse  studded  with  yellow  granules.  Conidiophores  100-300  by  3.5;£, 
arising  separately  or  from  portions  of  hyphse  just  above  the  surface  of  the  substratum. 
Conidial  fructification  50-100/i  in  length,  composed  of  one  verticil  of  branches  (some- 
times with  a  secondary  or  partial  secondary  verticil),  bearing  whorls  of  conidiiferous 
cells  11-12  by  2.5^,  narrowed  abruptly  to  form  sterigmata  at  the  apices.  Chains  long, 
divergent.  Conidia  elliptical,  3.4-3.8/i  by  2-2. 5/x,  green,  granular,  with  from  one  to 
several  small  highly  refractive  granules  in  each,  in  chains  falling  apart  in  fluid  mounts. 
Colonies  liquefy  sugar-gelatin  slowly  in  15  to  20  days. 

Received  from  Krai  in  Prague. 

The  authority  for  the  name  of  this  species  is  attributed  by  Saccardo 
to  Otto  Stoll,  as  here  indicated,  since  his  description,  or  rather  dis- 
cussion, of  this  fungus  forms  the  basis  of  Saccardo's  Latin  diagnosis. 
Stoll  quotes  the  name  from  Krai,  who  gives  the  author  as  Alex.  FleroflF, 
in  Warsaw.  Stoll  has  given  the  first  discussion  that  is  in  any  way 
adequate. 

A  closely  similar  organism  has  been  found  by  Prof.  F.  D.  Heald 
upon  corn  {Zea  mays)  in  Nebraska.  A  third  form  corresponding 
closely  in  morphology  and  many  cultural  characters  was  sent  from 
Miami,  Fla.,  by  Professor  Rolfs.  Although  distinguishable  by  some 
characters,  these  forms  resemble  P.  purpurogenum  as  described  above 
so  closely  in  morphology  and  cultural  characters  as  to  justify  inclu<ling 
them,  temporarily  at  least,  under  this  name.  Neither  of  these  forms 
produces  the  purple  color  as  rapidly  or  as  purely  as  the  original 
race  of  P.  purpurogenum. 

CULTURAL   DATA. 

Color  gray-green,  becoming  dark  green  with  the  presence  of  cane  sugar;  reverse 
yellow  to  reddish,  or  colorless;  color  in  media,  none  to  red  to  deep  purple,  almost  black, 
according  to  medium;  odor,  none. 

Fifteen  per  cent  gelatin  in  water,  medium  growth,  characteristic  fruiting;  liquefac- 
tion, partial  in  cultures  3  weeks  old,  or  none,  none  in  10  to  12  days;  litmus  reaction, 
slowly  alkaline  or  often  neutral.  Potato  agar  and  bean  agar,  good  colonies  but  no 
purple  color,  purple  produced  when  sugar  is  added.    Potato  plugs,  mycelium  yellow 


PENICILLIUM   PINOPHILUM. 


37 


with  granules,  potato  becomiug  purple.  Raulin's  fluid,  hIow  development.  Cohn's 
Bolution,  germination  only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar  10  per  cent  in  agar,  rich 
growth,  deep  red  medium.  Lactose  3  per  cent,  very  little  growth,  very  pale  purplish. 
Lactic  acid  0.9  per  cent,  medium  growth,  slowly  producing  purple  in  medium.  Levu- 
lose  3  per  cent,  small  colonies,  fluid  pale  red.  Galactose  3  per  cent,  small  colonies, 
acid  reaction.  Glycerin  3  per  cent,  no  growth.  Alcohol  3  to  5  per  cent  in  agar,  good 
growth  but  slow,  medium  becoming  purple.  Potato  starch  3  per  cent,  good  growth, 
abundant  purple  in  medium.  Tartaric  acid,  no  growth.  Butterfat,  very  slow  growth 
with  purple  color  in  fluid. 

Milk,  slow  development;  curdling  (0.25  per  cent  calcium  chlorid  added)  13  days; 
digestion  slow  and  only  partial;  color  in  milk,  shades  of  purple  according  to  progress, 
deepest  under  colony. 

Cooked  apple,  feeble  growth. 

Grew  equally  well  at  37°  C.  and  20°  C. 


Fig.  6. — Penidllium  purpurogcnum  O.  Stoll:  a,  b,  c,  conidial  fructification  showing  branching  and  arrange- 
ment (X  900);  o,  form  with  partial  secondary  verticil;  d,  e,f,  g,  conidliferous  cells  and  conidia  (X  1,400); 
li,},  k,  I,  m,  sketches  of  whole  fructifications  (X  140). 


PENICILLIUM  PINOPHILUM  Hedgcock  (nomen  novum). 

Syn.  Penidllium  aureum  Corda,  emended  Hedgcock,  Mo.  Bot.  Gard.  Rept.  17, 
pp.  105-107,  pi.  ii,  figs.  1-3. 
Not  P.  aureum  Corda,  Prachtflora,  p.  38,  t.  XVIII. 

Colonies  on  potato  or  bean  agar  and  milk  sugar  gelatin,  from  green  on  agar  through 
shades  of  yellow-green  to  bright  yellow  and  orange  on  media  containing  starch  and  cane 
sugar.  Superficial  hyphse  studded  with  yellow  granules  upon  acidfied  media.  Reverse 
of  colony  and  substratum  (upon  these  media)  colored  deep  rich  red.  Surface  growth 
partly  of  simple  conidiophores,  partly  aerial  hyphse,  and  ropes  of  hyphse  (which  rarely 
become  vertical  coremia)  bearing  conidiophores  as  lateral  branches.  Conidiophores 
100-2G0fi.  Conidial  fructifications  up  to  120fi  in  length,  consisting  of  single  verticils 
of  branches  10-16  by  2-2. 5/t,  bearing  whorls  of  conidliferous  cells  13-15  by  2-2.5/x  taper- 
ing into  acuminate  sterigmata  bearing  conidial  chains  which  are  parallel  but  do  not 
form  a  column.    Conidia  elliptical,  3-3.6  by  2[i,  smooth,  pale  green  or  yellowish  green. 


38 


CULTUEAL  STUDIES  OP  SPECIES  OP  PENICILLIUM. 


Colonies  liquefy  gelatin,  but  slowly  and  incompletely,  and  give  a  neutral  or  acid 
reaction  upon  all  litmus  media.  Under  different  conditions  of  culture  and  acidity  the 
discoloration  of  the  medium  varies  from  yellow  to  orange  and  deep  red.  Produces  dis- 
colorations  up>on  commercial  timbers.  Habitat,  pine  wood,  which  is  strongly  colored 
by  it. 

Culture  received  from  the  author,  G.  G.  Hedgcock,  of  the  Forest  Pathological  Lab- 
oratory, Bureau  of  Plant  Industry,  United  States  Department  of  Agriculture. 

Since  the  publication  of  his  description  of  this  fungus  (1906)  Hedg- 
cock® has  reached  the  conclusion,  concurred  in  by  the  Avriter,  that 
tliis  species  can  not  be  regarded  as  identical  with  the  species  described 
by  Corda  as  P.  aureum.  lie  notes  that  this  species  is  a  common  agent 
in  the  discoloration  of  pine  wood,  hence  proposes  the  name  P.  jyino- 
philum  (here  first  published).  Careful  consideration  of  Corda's 
figure  and  description  would  establish  a  strong  presumption  that 


Fig.  6.—PeniciUium  pinophilum  Hedgcock:  a,  young  conidial  fructiflcation  showing  conidiiferous  cells  at 
apex  of  central  branch  before  all  the  branches  appear  (X  1,600);  6,  a  verticil  of  four  branches,  upon  one 
of  which  fruit  appears  (X  1,600);  c,  d,  conidiiferous  cells  and  conidia  (X  1,600);  e,  rope  of  hyphoe  bearing 
conidiophores  sketched  (X  140);  /,  g,  forms  of  conidial  fructiflcation  (X  140). 

the  form  described  by  him  would  not  now  be  considered  a  species  of 
Penicillium. 

CULTURAL  DATA. 

Color,  conidial  areas  green,  vegetative  mycelium  colorless  or  studded  with  yellow 
granules;  reverse  of  colony  red;  color  in  media,  red. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  growth  slow,  surface  growth  of  conidiophores 
and  green  conidial  fructifications  only;  liquefaction,  none  or  very  slow  (in  acidified 
cultures  only  after  several  weeks);  litmus  reaction,  acid.  Potato  agar  and  bean 
agar,  mycelium  studded  with  yellow  granules,  conidial  areas  strict,  green;  reverse 
of  colony,  red.  Potato  plugs,  poor  growth,  not  typical.  Cohn's  solution,  spores 
germinated  only. 


PENICILLIUM  RUBRUM   AND  P.   LtJTEUM.  3d 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  growth  fair,  hyphae  yellow 
with  granules,  acid  reaction,  conidia  green,  reverse,  red.  Lactose,  growth  slow,  not 
characteristic.  Levulose  3  per  cent,  fair  growth,  yellow  mycelium,  acid  to  litmus. 
Galacto.se  3  per  cent,  fair  growth,  acid  reaction.  Glycerin,  very  slight  growth  which 
became  typical  when  sugar  was  added.  Potato  starch,  good  characteristic  growth. 
Butterfat,  slow  weak  growth,  with  characteristic;  colors  and  red  fluid. 

Milk,  growth  small;  curdling  (0.25  per  cent  calcium  chlorid  added),  very  slow; 
digestion,  none  or  very  slight;  color  in  milk,  red  at  top. 

At  37°  C,  grew  well,  more  rapidly  than  at  20°  C. 

PENICILLIUM  RUBRUM  O.  Stoll. 

Beitr.  z.  morp.  u.  biol.  char.  Penicillium,  Wiirzburg,  1904,  p.  35,  t.  I,  fig.  7;  t.  Ill, 
fig.  3;  t.  IV,  fig.  4. 

Colonies  upon  lactose  gelatin  and  potato  or  bean  agar,  from  green  through  ochraceous 
to  ochraceous  red  with  varying  conditions;  consisting  of  green  conidia  with  yellow 
mycelium  when  cane  sugar  is  added.  Aerial  portion  velvety  strict  or  very  closely 
floccose  in  media  without  sugar,  becoming  dense  cushions  of  mycelium  bearing  suc- 
cessive crops  of  green  conidia  in  cane-sugar  media;  marginal  growth  continuous  but 
slow  and  not  marked  by  a  white  border.  Reverse  and  mycelium  yellowish  to  red, 
yellow  in  sugar  media;  the  substratum  also  colored  in  old  agar  colonies.  Conidio- 
phores  arising  from  substratum  directly  or  as  very  short  lateral  branches  of  the  felted 
hyphae,  mostly  15-30  by  3-3.5/*,  swollen  at  the  apex,  making  a  dense  layer  on  the 
surface.  Conidial  fructification  usually  massed  into  a  heavy  column  with  a  broad 
triangular  base,  100-200/t  in  length,  from  a  dense  verticil  of  branches  of  the  conidio- 
phore,  each  swollen  at  the  apex.  Conidiiferous  cells  10-13  by  2-3«,  with  rather  abrupt 
points  from  which  the  conidia  are  cut  off.  Conidia  at  first  cylindrical,  then  elliptical 
or  even  globose,  3.4  by  2p.,  or  2.5-3. 3/i,  yellowish  green  to  dark  green  when  mature. 
Colonies  produce  slow  and  only  partial  liquefaction  of  sugar  gelatin. 

A  slow-growing  fungus  fruiting  for  several  weeks  and  differing  greatly  in  colors  with 
slight  and  undefined  differences  in  the  conditions.  Sometimes  producing  a  blood- 
red  color  on  the  reverse  of  the  colony. 

Cultures  received  from  Krai.     So  far  not  found  native  in  America. 

PENICILLIUM  LUTEUM  Zukal. 

Sitzber.  K.  Akad.  Wiss.  (Vienna)  Math.  Naturw.  Kl.,  XCVIII,  p.  521,  1889. 

Conidial  form:  Colonies  on  sugar  gelatin  and  yjotato  or  bean  agar,  white  or  gray 
or  transiently  yellow  on  media  lacking  sugars,  with  sometimes  greenish  areas  of  coni- 
dial fructification  showing  shades  of  yellow  (egg  yellow)  upon  sugar  media,  later 
passing  over  to  reddish,  especially  with  the  formation  of  aerial  wefts  or  balls  of  hyphae 
producing  asci  (several  weeks);  surface  rather  close  floccose,  spreading  indefinitely 
upon  the  substratum.  Reverse  of  colony  more  or  less  reddish,  especially  on  sugar 
media.  Conidiophores  a  thin  and  incomplete  layer,  scantily  produced  mostly  as 
lateral  branches  of  aerial  hyphae,  20-lOOjU  (mostly  30-60)  by  3/i.  Conidial  fructifica- 
tion usually  small  up  to  80/i  in  length,  commonly  with  a  single  lateral  branch  and  but 
two  verticils  of  long  acuminate  basidia  13-16  by  3^/(.  Conidia  elliptical  to  fusiform, 
2.4  by  2.3/t,  rather  firm  walled,  greenish,  swelling  greatly  and  producing  1  or  2  tubes 
in  germinating. 

This  species  characteristically  produces  yellow  mycelium,  from  which,  in  a  time 
varying  from  a  few  days  upon  media  rich  in  sugar  to  several  months  upon  plain  potato 
agar,  ascigerous  wefts  of  hyphae  arise.  As  given  by  Wehmer,-^  "ascigerous  concep- 
tacles"  are  0.5-2  mm.  in  diameter,  globose,  vitelline  then  red;  asci  reddish,  globose 
to  fusiform  8.8  by  7-7.8/e;  sporidia  4.8  by  3.3/i,  transversely  tricostate,  hyaline  to 


40 


CULTURAL  STUDIES  OF   SPECIES  OF   PENICILLIUM. 


reddish.  Colonies  do  not  liquefy,  or  only  slowly  and  partially  liquefy,  sugar  gelatin. 
Litmus  reaction  neutral,  the  shades  of  purple  found  aboHt  the  turning  point  of  litmus. 
Colonies  on  apple  are  brighter  yellow  and  produce  ascigerous  masses  in  very  few  days. 
Received  from  Professor  Thaxter,  identified  from  the  ascigerous  form  by  Dr.  C. 
Wehmer. 

The  author  is  convinced  that  Wehmer^^  is  in  error  in  attributing 
a  large  degree  of  polymorphism  to  this  fungus.  Numerous  cultures 
watched  under  all  sorts  of  conditions  are  evidence  that  this  species 
does  not  produce  prominent  coremia  or  large  masses  of  green  conidia. 

CULTURAL   DATA. 

Color  white,  but  mycelium  studded  with  yellow  granules  in  acid  media,  with  some- 
times reddish  areas  and  conidial  green  areas;  reverse  of  colony  yellow  to  orange; 
color  in  media,  pink  in  potato. 

Odor,  none. 


^=]3 


Fig.  7.—PeniciUium  rubrum  O.  Stoll:  a,  b,  c,  d,  e,  Whole  conidiophores  and  the  hranehing  of  conidial 
fiructifications  (X  1,400);  /,  j,  eonidiiferous cells  and  conidial  formation  ( X  1,400);  A,  j,  sketch  and  diagram 
of  habit  of  growth  (X  140);  t,  sketches  of  old  conidial  fractification  in  large  size(X  140);  rn,  diagrammatic 
figure  (the  successive  series  of  conidial  fructifications  are  produced  by  new  branches  from  hyphse  over- 
growing the  earlier  series).  (Drawn  from  gelatin  culture,  but  found  with  approximately  the  same 
morphology  upon  potato  agar.) 

Fifteen  per  cent  gelatin  in  water,  medium  growth;  liquefaction  none;  litmus 
reaction  acid;  potato  agar  and  bean  agar,  mycelium  transiently  yellow,  then  color- 
less or  reddish  or  yellowish  gray;  potato  plugs,  white  to  gray,  in  parts  yellow,  potato 
pinkish;  Raulin's  fluid,  slow  development,  bright  yellow  colonies;  Cohn's  solution, 
germination  only. 

Synthetic  fluid  (Box's),  carbon  supplied  as:  Cane  sugar,  good  growth,  yellow,  up 
to  30  per  cent  sugar,  no  growth  at  50  per  cent.  Lactose  3  per  cent,  very  slight  growth; 
lactic  acid  0.9  per  cent,  very  slight  growth.  Levnilose  3  per  cent,  very  slight  growth. 
Galactose  3  per  cent,  typical,  acid  reaction.    Glycerin  3  p>er  cent,  germination  only. 


t»ENICrLLIUM:  LUfEUM. 


41 


Fig  S.—Penicmium  luteum  Zukal:  a,  whole  conidiophore  and  fructification  (x  900);  6,  c,  d,  conidiiferous 
culls,  conidia  and  their  arrangement  (X  900);  g,  ft,  j,  k,  I,  m,  n,  sketches  of  fructifications  ( X  140);  o,  r, 
swelling  and  germination  of  conidia,  swollen  conidia  in  the  same  chain  with  those  which  refuse  to  grow 
(X  900). 


42 


CULTURAL  STUDIES  OF  SPECIES  OP  PENICILLIUM. 


Potato  starch  '.\  per  cent,  slow  developnieiit  but  characteristic  lUitterfat,  little  or 
no  growth . 

Cooked  apple,  mycelium  yellow,  ascigerous  maasee  produced  most  quickly  (1  week). 

Milk,  grows  poorly;  curdling  (with  0.25  per  cent  calcium  chlorid)  none;  diges- 
tion slight  and  slow.* 

At  37°  C,  grew  well;  at  20°  C,  more  slowly. 

PENICILLIUM  DUCLAUXI  Delacroix. 

Bulletin  de  la  Soci6t6  Mycologique  de  France,  Tome  VIII,  1891,  p.  107,  PI.  VII. 

Colonies  grown  upon  gelatin  and  potato  or  bean  agar  clear  dark  green  to  olive  when 
old,  consisting  of  short  crowded  conidiophores  arising  for  the  most  part  singly  from 
the  substratum  (strict),  but  sometimes  producing  short  coremia.  Long  coremia  are 
produced  abundantly  upon  orange,  milk,  potato,  and  all  media  rich  in  cane  sugar. 
Conidiophores  very  short,  10-50/i,  either  arising  directly  from  the  substratum  or  borne 


II  '*' 


Fig.  9.—Penicillium  duclauzi  Delacroix:  a,  b,  conldial  fructifications  with  young  conidia  smooth,  from 
IX)tato-agar  plate  culture,  simpler  types  (X  900);  c,  d,  e,  conidial  fructifications  from  potato-agar  plate 
culture,  more  complex  types  (X  1,400);  /,  g,  h,j,  sketches  of  habit  upon  potato  agar,  showing  the  very 
short  conidiophores  arising  from  the  substratum  ( X  140);  fc,  ripe  spores  highly  magnified  to  show  delicate 
markings  (X  900,  apochromatic);  I,  m,  n,  germination  of  conidia  (X  900,  apochromatic);  s,  t,  coremia, 
(sketch). 

upon  the  upper  third  of  the  coremia,  1-2  septate,  bearing  a  simple  conidial  fructifica- 
tion or  a  terminal  fructification  and  a  divergent  lateral  branch  with  a  whorl  of  coni- 
diiferous  cells.  Conidial  fructification  often  100-160/iin  length  consisting  of  a  fewconi- 
diiferous  cells  lO-12/i  in.  length  in  a  simple  terminal  whorl  or  less  commonly  in 
secondary  whorls.  Conidia  elliptical  fusiform  3.6-4  by  2-2.5//,  clear  homogeneous  green, 
smooth  when  young,  but  rugulose  when  ripe.  Colonies  liquefy  sugar-gelatin  in  Petri- 
dish  culture  slowly  from  twelfth  to  twenty-fifth  day  and  change  red  litmus  to  blue  in  7 
days.  Produces  a  coloring  agent  in  sugar  media  which  is  wine  red  in  alkaline  media 
and  yellow  (bile-yellow)  in  acid  media  (acts  as  an  indicator  with  neutral  point  very 
near  that  of  phenolphthalein). 

This  fungus  is  characterized  by  its  enormous  development  of  coremia  upon  milk, 
orange,  apple,  and  media  containing  cane  sugar,  while  producing  only  very  short 
conidiophores  in  bean  or  potato  agar  and  gelatin  free  from  sugar. 

Received  from  the  author,  George  Delacroix,  Paris,  but  one  culture  previously 
obtained  from  P.  H.  Rolfs  in  Florida  in  the  summer  of  1905  waa  thought  to  be  this. 


PENICILLIUM  CLAVIFORME. 


43 


This  species  has  not  been  found  commonly  in  America,  but  it  has 
been  many  times  observed  and  recognized  as  a  contamination  of 
other  cultures  in  this  laboratory  since  its  introduction.  Its  identi- 
fication is  therefore  easy. 

CULTURAL  DATA. 

Color,  fruiting  areas  olive  to  clear  deep  green,  often  brown  when  old;  reverse,  from 
colorless  to  yellow  and  shades  of  red,  according  to  medium;  color  in  media,  none  in 
media  free  from  sugar,  in  sugar  yellow  at  first,  then  slowly  red  (color  acta  as  an  indi- 
cator— acid  when  yellow,  alkaline  when  red). 

Odor,  rather  strong  in  Raulin's  fluid,  none  in  most  cultures. 

Fifteen  per  cent  gelatin  in  water,  medium  growth;  liquefaction,  none  in  15  days, 
then  slow  in  acidified  cultures  especially;  litmus  reaction,  neutral  slowly  or  weakly 
alkaline.     Potato  and  bean  agar,  green  fruiting  surfaces  consisting  of  very  short 
crowded    conidiophores    slightly 
tuberculate  at  times,  but  no  co- 
remia.     Potato    plugs,    abundant 
coremia,  deep  green,  reverse  and 
potato  yellowish.     Raulin's  fluid, 
good  growth,  many  coremia,  some 
odor.     Cohn's  solution,    germina- 
tion only. 

Synthetic  fluid  (Dox's),  carbon 
supplied  as:  Cane  sugar  3  per  cent, 
rich  growth,  coremia,  poor  colonies 
at  30  per  cent  and  60  per  cent. 
Lactose  3  per  cent,  very  weak 
growth,  few  very  small  coremia. 
Lactic  acid  0.9  per  cent,  food 
growth,  typical,  coremia  many, 
remains  acid.  Levulose  3  per 
cent,  weak  colonies.  Galactose 
3  per  cent,  poor  growth .  Glycerin 
3  per  cent,  germination  and  slow, 
weak  colonies.  Alcohol,  some 
growth.  Potato  starch,  rich 
growth,  coremia,  reverse  and  fluid 
yellowish  Butterfat,  slow  but 
typical  coremiform  colony,  fluid 
yellow  (acid  reaction). 

Milk,  typical  coremiform  colonies;  curdling  (0.25  per  cent  calcium  chlorid  added) 
in  9  days;  digestion  slow;  color,  becoming  yellow  (acid)  and  later  red  (alkaline)  in 
very  old  cultures. 

At  37*'  C.  no  growth;  check  grew  at  20°  C. 

PENICILLIUM  CLAVIFORME  Bainier. 

Bulletin  Trimestriel  de  la  Soci6t6  Mycologique  de  France,  Tome  XXI,  1905,  p.  127, 
PI.  XI,  figs.  8-11;  Saccardo,  Sylloge  Fungorum,  Vol.  XVIII,  p.  520. 

Colonies  on  milk-sugar  gelatin  and  potato  agar,  white  or  gray,  with  surface  com- 
posed of  loosely  floccose  hypha>,  bearing  simple  but  definitely  penicillate  fructifica- 
tions, between  the  bases  of  white  or  yellowish  simple  or  variously  branched  coremia 
1-2  cm.  long,  fertile  only  at  the  apices.  Simple  fructifications  sparingly  branched, 
bearing  small  verticils  of  conidiiferous  cells  9-10  by  2fi.  Coremial  fructifications 
consisting  of  closely  branching  and  interwoven  hyphse,  producing  verticils  of  conidiifer- 


FiG.  10. — Penicillium  daviforme  Bainier:  a,  coremium  grown 
upon  sugar  media,  showing  branching  stalk  with  several 
small  heads  of  conidia;  b,  coremium  grown  upon  gelatin 
free  from  sugar,  showing  typical  unbranched  stalk  with  a 
single  conidial  mass  splitting  as  it  increases  in  size  into 
several  columns  composed  of  chains  of  conidia.  (For  full 
illustration  of  the  structure  of  this  species,  see  Bainier's 
figures.) 


44  CULTURAL  STUDIES   OP   SPECIES  OF  PENICILLIUM. 

oiis  cells  crowded  into  a  false  hymcnium  and  producing  chains  of  conidia  adhering  in 
olive-green  masses  1-3  mm.  in  height.  Conidia  elliptical,  showing  a  connective, 
4.2^.6  by  3-3.3  fi,  homogeneous  green,  remaining  in  chains  in  fluid  mounts.  Colonies 
only  partially  liquefy  gelatin  media  and  give  a  weak  alkaline  or  neutral  reaction  with 
litmus. 
Received  from  G.  Bainier  October,  1905. 

A  culture  received  from  Reddick,  Ithaca,  N.  Y.,  marked  Whetzel 
No.  2095,  proved  to  be  this  species.  It  was  found  at  Junius,  N.  Y. 
A  culture  sent  to  Dr.  C.  H.  Peck  was  not  recognized.  Although  not 
closely  resembling  other  species  of  Penicillium  it  may  best  continue 
under  the  name  given  by  Bainier  until  closer  affinities  are  found  for  it. 

CULTURAL   DATA. 

Color,  mycelium  white  or  gray,  conidial  heads  olive  green;  reverse  of  colony  brown; 
color  in  media  brownish. 

Odor,  perceptible  in  media  containing  cane  sugar,  characteristic. 

Fifteen  per  cent  gelatin,  growth,  characteristic  coremia,  but  mainly  unbranched; 
liquefaction  slow,  partial;  litmus  neutral.  Potato  agar  and  bean  agar,  typical 
coremiform  colonies.  Potato  plugs,  vigorous  typical  growth  with  long  coremia. 
Raulin's  fluid,  characteristic.     Cohn's  solution,  characteristic. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  typical  growth  with  acid 
reaction  in  concentrations  from  1.5  to  30  per  cent,  coremia  branching,  no  coremia  at 
50  per  cent.  Lactose  3  per  cent,  slight  growth,  poorly  nourished  colonies,  very 
delicate  coremia.  Lactic  acid  0.9  per  cent,  germination  only.  Levulose  3  per  cent, 
slow-growing,  poorly  nourished  colony.  Galactose  3  per  cent,  typical  growth,  acid 
reaction.  Glycerin  3  per  cent,  not  typical,  no  coremia,  coremia  produced  after 
addition  of  cane  sugar.  Malic  acid  1  per  cent,  slight  growth,  few  and  very  small 
coremia.     Butterfat,  typical  colonies. 

Milk,  good  characteristic  growth;  curdling  (0.25  per  cent  calcium  chlorid  added) 
in  9  days;  digestion  slow  but  fairly  complete;  color  in  milk  brown  or  reddish. 

At  37°  C,  no  growth  in  6  days;  culture  grew  when  cooled  to  20° C. 

PENICILLIUM  GRANULATUM  Bainier. 

Bui.  Soc.  MycoL  France,  XXI,  1905,  p.  127,  PI.  XI,  figs.  6,  7. 

Colonies  upon  plain  gelatin  and  potato  or  bean  agar  yellowish  green  to  gray  or 
grayish  brown^  superficially  composed  of  crowded  small  coremia  1-3  mm.  in  height, 
mixed  with  floccose  hyphae  and  separate  conidiophores,  spreading  indeterminately 
upon  the  substratum.  Reverse  reddish  orange  (approaching  "fulvous"),  aerial 
hyphae  delicately  granular  or  spinulose,  which  separates  this  from  all  other  species 
studied.  Conidiophores  4-4. 5/z  in  diameter,  short  or  very  long,  either  separate  or, 
mostly,  massed  into  very  short,  crowded  coremia  (less  than  1  mm.  in  height).  Conidial 
fructifications  usually  100-200/x  in  length,  once  or  twice  verticillate,  with  many 
conidiiferous  cells  9  by  2-2.5;x,  and  long,  loosely  divergent  chains  of  conidia.  Conidia 
at  first  cylindrical,  then  elliptical  to  globose,  about  2.5-3  by  3-3.5  or  3/i  in  diam- 
eter, yellowish  green,  granular,  remaining  in  long  chains  in  fluid  mounts.  Colonies 
do  not  liquefy  gelatin,  litmus  reaction  slowly  alkaline. 

The  delicately  granular  or  spinulose  hyphae  as  noted  and  figured 
by  Bainier  are  a  valid  and  distinctive  character.  The  species  is  also 
easily  recognized  by  its  general  appearance  and  habit.  Obtained 
from  the  type  cultures  of  Professor  Bainier,  Paris. 


PENICILLIUM   BREVICAULE. 


45 


CULTURAL  DATA. 

Color  yellowish  green  to  green  and  later  various  shades  of  brown;  reverse  of  colony 
orange,  or  yellow  to  deep  orange,  or  even  red;  color  in  media  yellow  to  orange  to 
red  in  media  containing  starch  or  sugar. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  characteristic  growth;  liquefaction,  none,  or 
partial  after  several  weeks  in  acidified  cultures;  litmus  reaction,  strongly  alkaline. 
Potato  agar  and  bean  agar,  yellow-green  to  brown,  yellow  below,  coremia  not  closely 
crowded  as  in  sugar  media.  Potato  plugs,  characteristic  growth,  potato  stained  yel- 
low to  deep  brownish  yellow.  Raulin's  fluid,  characteristic  growth.  Cohn's  solution, 
germination  only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  3-30  per  cent,  characteristic 
growth  and  colors,  but  not  normal  at  60  per  cent,  with  acid  reaction.  Lactose  3  per 
cent,  characteristic  structure  but  weak  development.  Lactic  acid  0.9  per  cent, 
growth,  but  not  normal  colonies.  Levulose  3  per  cent,  slow  development,  small  colo- 
nies. Galactose  3  per  cent,  characteristic.  Glycerin  3  per  cent,  slight  growth,  grew 
freely  when  cane  sugar  was  added.  Potato  starch,  characteristic.  Butterfat,  typical 
growth,  reverse  and  fluid  orange;  yellowish. 


Fig.  11. — PeniciUium  grantUatum  Balnier:  a,  b,  c,  branching  of  conidlal  fructification  showing  grana- 
lated  echinulated  cell  walls  (X  900);  d,  e,  f,  g,  conidiiferous  cells  and  conidial  chains  (X  1,400);  ft,  j,  k, 
sketches  of  fructifications  (X  140);  m,  sketches  drawn  from  photomicrograph  of  coremia  showing  fertile 
and  sterile  areas  (X  about  25). 

Milk,  rapid  and  characteristic  growth;  curdling,  none;  digestion,  slow  but  complete 
(fairly);  color  in  milk,  yellow  to  orange  to  deep  red. 

At  37°  C,  no  growth,  conidia  grew  when  removed  to  lower  temperature;  at  20°  C, 
excellent  growth. 

PENICILLIUM  BREVICAULE  Saccardo. 

Fungi  Italici,  No.  893,  Mich.,  II,  p.  547. 

Colonies  grown  upon  sugar  gelatin  grayish  white,  then  yellowish  brown  or  chocolate, 
consisting  of  short  closely  crowded  conidiophores  making  powdery  areas  overgrown  by 
loosely  trailing  floccose  hyphae  and  ropes  of  hyphae,  with  broadly  spreading  indeter- 
minate margin.  Coridiophores,  short,  10-30;i  mostly,  arising  directly  from  the  sub- 
merged hyphse,  or  numerously  and  irregularly  borne  as  lateral  and  perpendicular 
branches  of  trailing  aerial  hyphae  and  ropes  of  hyphae.  Conidial  fructifications  either 
simple  chains  terminating  unbranched  or  sparingly  branched  conidiophores  in  young 
colonies,  or  verticillately  and  irregularly  twice  verticillately  branching  systems  bear- 
ing numerous  divergent  chains  often  150^  in  length  in  old  colonies.    Conidiiferoue 


46 


CULTURAL  STUDIES  OF  SPECIES  OF  PENICILLIUM. 


cells  continuous  with  conidiophores  12-15  by  4/i  tapering  to  slender  sterigmata.  Co- 
nidia  somewhat  pear-shaped,  slightly  tuberculate  at  apex,  with  broad  base,  6.5-7.5  by 
7.5-9/i,  in  mass  light  brown  to  chocolate;  at  first  smooth,  then  with  thick  tuberculate 
walls,  viable  for  many  months,  germinating  by  a  single  tube  from  the  thin  center  of 
the  broad  base  into  a  bulbous  enlargement  from  which  mycelial  hyphae  about  2//  in 
diameter  arise.  Mycelium  very  thin  walled,  narrow  cells  of  varying  length.  Colonies 
liquefy  sugar  gelatin  and  give  a  strong  blue  reaction  in  litmus  media,  but  grow  very 
tardily,  if  at  all,  in  potato  or  bean  agar.  Grows  very  rapidly  upon  neutral  or  alkaline 
media,  but  very  slowly  or  not  at  all  in  media  acid  to  litmus.  Digests  milk.  Refused 
to  grow  after  repeated  inoculation  into  sterilized  apple.  Spores  which  refused  to 
germinate  in  the  agar  media  used  grew  immediately  when  transferred  to  any  of  the 
gelatin  media. 

Cosmopolitan,  forms  characteristic  chocolate  patches  on  Camembert  cheese.     Se- 
cured from  numerous  brands  of  cheese  and  common  in  the  laboratories  of  this  station. 


•■^rs 


Fig.  12.— Penicillium  brcvicaule  Saccardo:  a,  conidiophores  and  simple  conidial  chains  with  spores  still 
smooth  (X  900);  b,f,  more  complex  conidial  fructifications  (X  900);  c,  tv.-o  yoiing  conidial  chains,  show- 
ing thick  walls  of  spores  (X  1,400);  d,  c,  eonidia  after  becoming  pchinulato  (X  1,400);  g,  h,j,  sketches  of 
forms  and  habit  of  conidial  fructifications  (X  liO);'g  from  an  old  eluture,  sessile  or  almost  so;  h  andj 
show  trailing  hyphae  and  a  rope  of  hyphse  with  lateral  conidiophores;  k,  germinated  conidium  where 
the  old  spore  wall  lies  empty  beside  the  growing  cell  (X  1,400). 

The  author  does  not  believe  that  this  species  is  closely  related  to 
other  species  of  this  genus,  but  since  it  has  been  placed  here  by  a 
very  liberal  interpretation  of  descriptions  it  may  perhaps  remain 
under  tliis  name  until  someone  finds  out  its  real  affinities.  The  two 
forms  which  follow  as  varieties  are  found  in  the  same  habitat,  show 
closely  similar  morphology,  and  give  almost  identical  physiological 
reactions.  Their  designation  as  varieties  of  P.  hrevicaule  may  there- 
fore be  justified,  though  one  at  least  (var.  glabrum)  seems  separate 
enough  to  warrant  proposing  for  it  a  specific  name. 

CULTURAL  DATA. 

Color,  conidial  surfaces  clay-yellow  to  chocolate;  reverse  of  colony,  mycelium 
colorless;  color  in  media,  none. 
Odor,  ammoniacal,  used  as  a  test  for  arsenic  (Gosio  et  al.). 


PENICILLIUM   BREVICAULB. 


47 


Fifteen  per  cent  gelatin  in  water,  typical  colony;  liquefaction,  rapid — 5-6  days; 
litmus,  strongly  alkaline.  Prefers  alkaline,  neutral,  or  only  slightly  acid  media. 
Potato  agar,  spores  sometimes  refuse  even  to  germinate,  but  grow  when  transferred 
to  gelatin;  grows  poorly  on  agar  media  free  from  sugar,  or  peptone,  or  the  by-products 
of  other  fungus  growth .  Common  as  a  secondary  growth  in  such  plates.  Potato  plugs, 
good  colonies.  Cohn's  solution,  characteristic  but  slow  growth.  Raulin's  fluid, 
germinated,  but  very  slight  growth. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar  in  agar  3  per  cent,  failed 
to  grow,  same  spores  transferred  to  gelatin  media  grew  promptly.  Lactose  10  per  cent, 
slow,  characteristic  color  to  conidia.  Lactic  acid  0.9  per  cent,  germination  only. 
Levulose  2.5  per  cent,  growth  characteristic,  alkaline  reaction  to  litmus.  Galactose 
3  per  cent,  characteristic  growth,  alkaline  reaction  to  litmus.  Glycerin  3  per  cent, 
spores  germinated  only,  grew  well  upon  addition  of  cane  sugar.  Potato  starch  grew 
well.  Malic  acid,  germinated  only.  Butter  fat,  slow  development,  finally  producing 
drops  of  yellow  oil  which  separate  out. 

Milk,  rapid  growth;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  10  days; 
digestion,  rapid;  color  in  milk,  none. 

Cooked  apple,  failed  to  grow  after  repeated  inoculation. 

At  37"  and  20°  C,  grew  equally  well. 


Fig.  13. — Penicillum  brevicaule,  var.  album:  a,  h,  c,  d,  conldiophores  and  fructification  borne  variously  and 
differently  branching  (X  900);  e,/,rii)6  conidia;  j?,  ft,  slictch  of  single  hyphae  and  a  rope  of  hypha?  bearing 
conldiophores  (X  140);  j,  germinated  spore. 

PENICILLIUM  BREVICAULE  Saccardo,  var.  ALBUM  Thorn,  n.  var. 

Colonies  upon  sugar  gelatin  white  to  cream-colored  alike  above  and  below  upon  all 
media,  strict  to  sparsely  floccose,  with  trailing  hyphse  and  ropes  of  hyphae,  indeter- 
minately spreading.  Conldiophores  either  arising  from  substratum  directly  or  mostly 
as  perpendicular  branches  of  aerial  hyphae  and  ropes  of  hyphae  15-40/i  in  length, 
conidial  fructification  varying  from  a  single  chain  to  more  or  less  complex  penicillate 
branching,  mostly  producing  few  chains  of  indefinite  length  and  arrangement  from 
narrow  tapering  conidiiferous  cells.  Conidia  pyriform  to  subglobose,  with  basal  collar, 
9-10/1,  roughly  tuberculate,  white  or  slightly  yellowish  tinged,  thick  walled  except  at 
the  base,  the  center  of  which  remains  as  a  germ  pore.  Colonies  rapidly  liquefy  sugar 
gelatin  with  strong  ammoniacal  odor,  and  give  an  intensely  alkaline  reaction  in  litmus 
media.  Gives  exactly  the  same  reactions  as  P.  brevicaule  Sacc.  Differs  from  the 
latter  slightly,  except  in  the  color  of  the  spores. 

Common  upon  imported  Camembert  cheese.  Found  often  upon  domestic  Camem- 
bert  and  grows  very  readily  in  cheese  cellars,  where  it  becomes  a  nuisance. 


48 


CULTURAL  STUDIES  OF   SPECIES  OP  PENICILLIUM. 


CULTURAL  DATA. 

Same  as  P.  hrevicaule  Sacc,  except  the  following  difference  noted: 

Conidia  cream,  somewhat  larger  than  P.  hrevicaule. 

Cohn's  solution  failed  to  produce  characteristic  colonies. 

In  Dox's  solution,  with  butterfat  as  a  source  of  carbon,  it  differs  from  P.  hrevicaule  by 
failing  to  cause  drops  of  yellow  oil  to  separate  out. 

Agar-agar:  In  repeated  cultures  this  organism  has  failed  to  grow  well  in  agar  media. 
In  some  such  cases  the  spores  transferred  from  the  agar  to  gelatin  grew  at  once.  Some 
cultures  upon  agar  grow  slowly  and  in  typical  manner,  but  the  development  upon  all 
agar  media  has  seemed  uncertain.  In  syitfhetic  solution  cultures  were  obtained  when 
the  inoculation  of  tubes  of  the  same  solution  with  1.5  per  cent  of  agar  added  to  make  a 
solid  substratum  produced  no  growth. 

PENICILLIUM  BREVICAULE  Saccardo,  var.  GLABRUM  Thorn,  n.  var. 

Colonies  white  or  only  slightly  yellowish-tinged  in  all  gelatin  media,  grow  not  at  all 
or  with  difficulty  on  agar  of  most  formulae.  Aerial  portion  consisting  of  short,  closely 
crowded  conidiophores  making  a  powdery  surface  overgrown  by  loosely  trailing 
hyphae  and  ropes  of  hyphae,  spreading  broadly  over  the  substratum.  Conidiophores, 
short,  mostly  10-30/t,  arising  directly  from  submerged  hyphae  or  numerously  and  irregu- 


FiG.  14.— Pen»e»/Zi«m  hrevicaule,  var.  glabrum:  a,  b,  c,  branching  of  conidial  fructification  (X  900);  d,  chain 
of  conidia  (X  1,400);  /,  formation  of  conidium  on  young  branch  (X  1,400);  g,  h,  sketch  of  appearance 
in  culture  (X  140);  m,  n,  o,  germination  of  conidia. 


larly  borne  as  perpendicular  branches  of  the  superficial  hyphae  and  ropes  of  hyphae. 
Conidial  fructifications  from  simple  chains  of  spores  to  fairly  complex  penicillate 
groups  of  branchlets  resembling  P.  hrevicaule,  but  mostly  less  complex.  Conidia 
obovate,  pyriform  7-8  by  8-10/t  or  almost  globose,  7-9/i,  smooth,  white,  rather  thick- 
walled  and  retaining  their  power  to  germinate  for  many  months.  In  old  potato  and 
other  cultures  black  sclerotia  are  formed  in  the  substratum  but  do  not  produce  asci. 
Liquefies  gelatin  rapidly  (within  one  week),  gives  a  strong  alkaline  reaction  and 
ammoniacal  odor. 

Habitat,  found  repeatedly  on  imported  Camembert  cheese  and  secondarily  upon 
domestic  soft  cheese,  where  it  grows  into  prominent  cottony  patches  indistinguishable 
to  the  eye  from  the  white  variety  of  P.  hrevicaule.  This  fungus  is  separated  from 
P.  hrevicaule  by  itd  smooth  white  spores  and  the  production  of  the  black  sclerotia  in 
the  substratum. 


PENICILLIUM  ROSEUM. 


49 


This  form  is  certainly  closely  related  to  P.  brevicaule  by  its  physiological  reactions 
and  its  general  morphology.  It  was  found  in  one  case  among  the  exsiccati  in  the 
Ilarvard  herbarium  under  the  name  of  Monilia  Candida.  There  is,  however,  no  possi- 
bility of  confusing  this  form  with  that  species  as  understood  and  described  by  more 
recent  students,  such  as  Hansen  and  Jorgenson. 

CULTURAL   DATA. 

Exactly  as  in  P.  brevicaule,  except  for  the  following: 
Conidia  smooth,  somewhat  smaller. 
Color  more  nearly  white. 

Sclerotia  black,  found  in  very  old  potato-plug  cultures  or  in  agar  cultures  which 
have  grown  several  weeks  or  months. 
Cohn's  solution  failed  to  produce  a  characteristic  colony. 

PENICILLIUM  ROSEUM  Link  (?). 

Colonies  on  milk-sugar  gelatin  or  potato  agar  white  to  pink  or  salmon  in  fruiting 
areas,  loose  floccose  with  simple  hyphae  and  ropes  of  hyphee,  producing  dense  irregular 


Fig.  15.— Penicillium  roseum  Link  (?):  o,  6,  c,  branching  of  conidial  fructification,  showing  few  colls  in 
each  verticil  ( X  900);  d,  e,  conidiiferous  cell  and  conidia  ( X  900);  g,  h,  j,  k,  sketches  of  ripe  fructifications 
showing  agglutination  of  conidia  into  slimy  masses  (X  140). 

pinkish  masses  or  sclerotia  up  to  1  mm.  or  more  in  diameter  in  old  culture.  Coni- 
diophores  borne  as  perpendicular  branches  of  aerial  hyphae  or  ropes  of  hyphae,  45-125/i. 
Conidial  fructification  up  to  140/x  in  length,  once  or  twice  irregularly  alternately  or 
verticillately  branched,  with  conidiiferous  cells  varying  from  12  by  2-3;u  in  the  verticils 
of  5  or  less  to  17  by  2.3/t  when  solitary,  bearing  conidia  which  become  aggregated  into 
gelatinous  balls  or  masses.  Conidia  colorless  (pink  or  rosy  in  mass),  elliptical,  5-7  by 
3-5;/,  slightly  apiculate,  smooth,  appearing  delicately  granular  within.  Colonies 
liquefy  gelatin  cultures  rapidly  and. give  an  alkaline  reaction  to  litmus  media. 
Brought  from  Krai,  in  Prague,  Bohemia. 
8108— Bull.  118—10 i 


50  CULTURAL  STUDIES  OF  SPECIES  OF  PENICILLIUM. 

The  same  organism  has  been  found  once  in  accidental  culture  in  tliis 
laboratory;  received  once  from  a  correspondent  in  Halle,  Germany, 
and  later  found  under  this  name  as  No.  1179  in  De  Thiimen's  Myco- 
theca  Universalis;  collected  by  Ravenel  in  South  Carolina  in  1876 
upon  leaves  of  Buxus;  this  and  several  other  specimens  were  found 
in  the  mycological  collection  of  the  Bureau  of  Plant  Industry,  United 
States  Department  of  Agriculture.  The  spores  are  the  same  length 
as  given  by  Saccardo,  but  slightly  broader.  The  number  of  specimens 
found  under  this  name  from  widely  different  workers  appears  to 
justify  the  belief  that  this  is  the  organism  described  by  Link  under 
this  name.  If  the  development  of  a  mucilaginous  mass  enveloping 
the  conidia  be  regarded  as  a  sufficient  basis  for  separation  of  such 
species  under  the  generic  name  of  Gliocladium,  this  species  would 
become  Gliocladium  roseum  (Link), 

The  form  upon  Buxus  is  cited  by  Saccardo,  referring  to  it  as  "P. 
roseum  Cooke,  non-Link,"  and  held  to  be  VerticiUium  huxi  Auersw, 
et  Fleisch.  Examination  of  the  material  would  indicate  that  in 
De  Thiimen's  collection  at  least  this  species  is  more  closely  allied  to 
the  other  species  of  PeniciUium  than  to  VerticiUium. 

CULTURAL  DATA. 

Color  white  or  shades  of  salmon  pink;  reverse  cream  or  white;  color  in  media, 
none. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  medium  growth,  white.  Liquefaction,  rapid. 
Litmus  reaction,  alkaline,  strongly.  Potato  agar  and  bean  agar,  good  growth,  but 
white.     Potato  plugs,  white  colonies.    Cohn's  solution,  slight  growth. 

In  Dox's  solution,  with  butterfat  as  a  source  of  carbon,  this  species  caused  drops 
of  yellow  oil  to  separate  out. 

At  37°  C,  killed;  check  at  20°  C,  good. 

PENICILLIUM  CAMEKBERTI  Thorn. 

Emended  from  U.  S.  Department  of  Agriculture,  Bureau  of  Animal  Industry,  Bui. 
82,  p.  33,  fig.  1,  1906. 

Possible  syn.:  P.  album  Epstein  (not  Preuss),  Archiv  f.  Ilyg.,  Bd.  45,  Hft.  4, 
p.  360,  1902. 
P.  epsteini  Lindau,  Deutschl.  Krypt.  Flora,  Pilze,  VIII,  p.  166. 

Colonies  on  potato  agar  or  lactose  gelatin  effused ;  white  (sometimes  yellowish  white), 
changing  in  5-8  days  to  gray -green  (glaucous);  surface  of  colony  floccose,  of  loosely 
felted  hyphse  about  5/t  in  diameter,  reverse  of  colony  yellowish  white;  conidiophores 
300-800;/  in  length,  3-4;i  in  diameter,  septate,  cells  thin-walled,  often  collapsing  in 
age,  arising  as  branches  of  aerial  hyphae;  fructification  sometimes  175/x  in  length,  but 
usually  much  less,  consisting  commonly  of  one  main  branch  and  one  lateral  branch, 
sparingly  branched  to  produce  rather  few  conidiiferous  cells  which  bear  long  loosely 
divergent  chains  of  conidia.  Conidiiferous  cells  8-11  by  2.4-3;t.  Conidia  at  first 
cylindrical,  then  elliptical,  and  finally  globose  when  ripe,  smooth  bluish  green  by 
transmitted  light,  thin-walled  and  commonly  guttulate,  4.5-5.5/z  in  diameter,  swelling 
in  germination  to  8-10/t.  Germ  tubes  one  to  several.  Cells  of  mycelium  about  5 
by  20-40/4.     Liquefies  lactose  gelatin  only  under  center  of  colony.     Produces  a  strong 


PENICILLIUM   CAMEMBERTI. 


51 


alkaline  reaction  in  gelatin,  free  from  sugar,  but  in  sugar  media  produces  a  more  or 
less  persistent  acid  reaction.     Growing  and  fruiting  period,  about  2  weeks.     Fruits 
only  upon  exposed  surfaces  of  the  substrata;  never  produces  spores  in  cavities  not 
broadly  open. 
Habitat,  Camembert  and  other  soft  cheeses. 

The  name  P.  album  Epstein,  which  is  also  P.  epsteini  Lindau/**  is 
inserted  in  the  list  of  possible  synonymy  because  this  name  is  accepted 
for  this  mold  by  Maz6  ^^  in  a  recent  paper.  If  we  base  identification 
upon  the  cultural  charac- 
ters given  for  his  mold 
by  Epstein,  it  could  not 
have  been  P.  camemberti. 
The  characterization,  so 
far  as  given  by  Epstein 
and  extracted  from  his 
paper  by  Lindau,  might 
refer  to  the  pure  white 
form  (P.  candidum)  as 
interpreted  by  Roger  and 
Maz6  better  than  to  this 
species.  There  is  cer- 
tainly at  least  a  varietal 
difference  (spore  color) 
between  P.  camemberti 
and  the  P.  candidum  of 
Roger  and  Maz6,  which 
is  designated  a  variety  of 
P.  camemberti  in  this 
paper. 

The  change  in  litmus 
reaction  in  this  descrip- 
tion from  that  previously 
published    is    result    of 
proof  that  no  acidity  is 
produced  by  this  species 
except  when  sugar  is  pres- 
ent.   A  change  from  blue 
to  red  and  black  to  blue 
indicates  the  production 
of  acidity  by  fermenta- 
tion of  sugar  followed  by  the  neutralization  of  the  acid  so  produced 
by  the  alkaline  by-products  of  proteolysis  or  by  a  further  change 
in  the  acid.     Therefore  this  statement  is  best  omitted  from  the  diag- 
nostic description. 


Fig.  16. — Penicillium  camemberti  Thom:  a,  conldiophore  show- 
ing a  common  type  of  branching  and  the  production  of  basida 
and  conldia,  highly  magnified;  6,  a  common  form  showing 
much  less  branching;  c,  d,  f,  diagrams  of  large  fructifications 
(X  80);  g,  i,j,  germinating  conidia.  (From  Bull.  82,  Bureau 
of  Animal  Industry.) 


52  CULTURAL  STUDIES  OP   SPECIES  OF  PENICILLIUM. 

CULTURAL  DATA. 

Color  white,  through  cream  to  a  shade  of  gray-green  upon  sugar  media,  without 
sugar  gray  or  drab,  all  becoming  drab  when  old  (exposure  to  light);  reverse  of  colony, 
uncolored,  cream;  media,  uncolored. 

Odor,  none,  except  in  very  old  cultures  upon  milk  or  cereal. 

Fifteen  per  cent  gelatin  in  water,  moderate  growth,  not  rich;  liquefaction,  none, 
or  slow  and  partial  after  2  to  3  weeks  in  acidified  cultures;  litmus,  strongly  alkaline; 
colonies  slow,  moderate  growth,  typical.  Potato  agar  and  bean  agar  and  potato  plugs, 
fair,  white  to  gray  colonies  lacking  or  nearly  free  from  green  color.  Raulin's  fluid, 
very  heavy  growth,  white  through  pronounced  cream  to  green  shades.  Cohn's  solu- 
tion, slow  growth,  charactoristic,  but  lacking  in  green  color. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  characteristic,  abundant 
green  color  in  solutions  as  high  as  50  per  cent,  acid.  Lactose  3  per  cent,  characteristic, 
acid  reaction.  Lactic  acid  0.9  per  cent,  slowly  typical,  with  ultimately  an  alkaline 
reaction  to  litmus.  Levulose  3  per  cent,  slowly  typical.  Galactose  3  per  cent,  char- 
acteristic growth,  very  slowly  developing  the  typical  green  color.  Alcohol  about  5 
per  cent,  characteristic.  Potato  starch,  characteristic.  Malic  acid  1  per  cent,  fair 
colony,  colorless.  Butterfat,  slowly  typical  colony  with  a  tinge  of  violet  in  reverse, 
no  color  in  fluid. 

Milk,  typical  growth;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  10  days; 
digestion,  slow  but  fairly  complete;  color  in  milk,  none. 

Camembert  cheese,  capable  of  reducing  moist  curd  soured  by  lactic  organisms  to  a 
semiliquid  condition. 

At  35°  to  37°  C,  no  growth;  at  20°  C,  excellent. 

PENICILLIUM  CAMEMBERTI,  var.  ROGERI  Thorn,  n.  var. 

Syn.  P.  candidum  of  Roger  and  Maze,  not  Link. 

Colonies  grown  upon  sugar  gelatin  or  bean  or  potato  agar  pure  white,  loosely  and 
evenly  floccose  to  the  very  margin,  where  aerial  and  submerged  hypha?  grow  with 
equal  rapidity.  Reverse  of  colonies  white  or  yellowish  white  (not  discolored).  Co- 
nidiophores  3-5  by  100-800/i  varying  greatly ,  mostly  branches  of  aerial  hyphse.  Conidial 
fructification  70-90/i  in  length,  loosely  and  irregularly  branched  and  bearing  rather 
few  basidia  at  unequal  heights,  with  divergent  chains  of  colorless  conidia.  Branch- 
ing system  of  conidial  fructification  sometimes  75/i  in  length.  Conidia  smooth, 
hyaline,  4-4.5  or  even  5.5//  in  diameter,  globose  or  nearly  so  when  ripe.  Sugar  gelatin 
is  slowly  liquefied  under  the  center  of  the  colony  only,  colonies  never  floating  in  a 
pool  of  liquid.  Reaction  in  the  medium  is  acid  to  litmus  at  first,  then  changes  to 
alkaline. 

This  fungus  has  been  found  by  the  author  only  upon  Camembert, 
Brie,  and  Neufch&tel  cheeses  from  western  Europe. 

This  variety  has  been  discussed  by  Maze  ^^  as  P.  candidum  Link. 
This  seems  an  impossible  application  of  the  name  P.  candidum  from 
Link's  description  "  or  that  given  by  Saccardo,  since  tlie  spores  are 
stated  to  be  2-3/z  in  diameter.  Further,  in  such  identifications  no 
account  is  taken  of  a  paper  by  Morini,*"  in  which  an  ascigerous  stage 
is  described  for  P.  candidum  Link.  Under  four  years  of  cultivation 
no  signs  of  an  ascigerous  form  have  been  produced.  Stoll  '*  has  consid- 
ered P.  candidum  to  be  only  a  colorless  P.  glaucum,  but  as  the  author 
has  so  far  failed  to  find  a  worker  who  will  undertake  to  limit  the  name 


PENICILLIUM  CAMEMBERTI. 


63 


P.  glaucum  to  a  special  form,  this  does  not  mend  matters.  Long 
cultivation  does  show,  however,  that  this  organism  is  closely  related 
to  the  one  already  described  as  P.  camemberti.'^  Since  this  is  the 
form  given  prominence  in  cheese  studies  by  the  work  of  Georges 
Roger,"  it  seems  most  natural  to  regard  it  as  a  variety  of  the  former 
species  and  designate  it  by  Roger's  name. 

In    Lafar's    Ilandbuch    der    Technischen    Mykologie,    Professors 
Weigmann  ^^  and  Wehmer  ^*  refer  to  this  fungus  as  probably  iden- 


Fio.  V.—Penicillium  camemberti,  var.  rogeri:  a,  b,  c,  types  of  branching  of  conidial  fructification  (X  900); 
e,  ripe  conidia,  showing  variation  in  size  (X  900);  g,  h,j,  k,  sketches  of  conidial  fructifications  (X  140); 
I,  m,  germination  of  conidia  by  several  tubes. 

tical  with  P.  camemberti.  However  this  may  appear  from  examina- 
tion of  the  literature  alone,  no  one  actually  familiar  with  the  cultures 
will  claim  such  identity.  If  the  possession  of  colorless  spores  be 
regarded  as  a  case  of  albinism,  this  form  may  perhaps  be  regarded 
as  a  variety  of  P.  camemberti.  It  has  been  kept  separate  and  re- 
mained constant  in  culture  for  several  years.  It  would  seem,  there- 
fore, equally  proper  to  regard  it  as  a  different  species  were  it  not  so 
closely  associated  with  P.  camemberti  in  every  other  character. 


64  CULTURAL  STUDIES   OF   SPECIES  OF   PENICILLIUM. 

The  name  P,  cajulidum  was  used  by  Link  for  a  mold  growing  upon 
decaying  leaves,  bulbs,  and  fungi,  which  was  said  to  be  common, 
and  Morini  "  later  describes  an  ascigerous  form  of  this  fungus.  The 
spores  are  of  different  size  (2-3;^).  There  appears  to  be  no  justifica- 
tion for  adopting  this  name  for  the  fungus  used  by  Roger  in  cheese 
making. 

CULTURAL   DATA. 

Different  from  P.  camemberti  only  as  follows: 
Color  of  conidia  persistently  white. 

Cohn's  solution,  failed  to  germinate.     Same  spores  transferred  to  gelatin  after  4 
month-s  grew  nonnally. 
Camembert  cheese,  does  not  produce  the  same  texture  as  the  preceding  species. 

PENICILLIUM  BIFORME  n.  sp. 

Latin  diagnosis."' — Coloniis  in  gelatina  cultis,  albis,  lente  glaucescentibus,  densius 
fioccosis,  margine  sterili  lata,  aut,  in  agaro  solani  tuberosi  cultis,  albis  glaucescentibus, 
mox  avellaneis  vel  fere  olivaceis,  parte  aeria  ex  conidiophoris  brevissimis  et  creberrimis 
fructibus  conidicisque  composita;  conidiophoris  (sine  ramis)  60-150/t  in  agaro,  vel 
longioribus  ramosis  ex  hyphis  fioccosis  in  gelatina  cultis;  fructibus  conidicis  100-200/t 
longis,  plerumque  1-2  ramosis  alternates,  ramis  convergentibus  vel  divergentibus, 
ramulis  verticillatisbasidiaapice  verticillatagerentibus;  basidiis  8-10  usque  13X3//, 
conidiis  primum  ellipticis  vel  cylindricis  demum  globosis,  3.2-3.5X4-4.3,  vel  4fi 
diam.,  in  catenis  manentibus  submersis;  coloniis  copiosis  in  saccharo  lactis,  gela- 
tinam  in  parte  lente  liquefacientibus,  alkalinis  lacmo,  odore  mucidis. 

Habitat,  in  caseo,  ex  Gallia. 

Cultivated  in  gelatin,  white,  slowly  gray-green,  densely  floccose,  with  broad  vegeta- 
tive margin,  spreading  widely  over  the  substratum;  in  potato  agar  white,  then  gray- 
green,  rapidly  becoming  yellowish-brown,  drab,  or  almost  olive,  restricted  in  growth, 
aerial  portion  consisting  of  very  short  densely  crowded  conidiophores  and  conidial 
fructifications;  conidiophores  60-150/x  on  agar  or  slightly  longer  when  arising  as 
branches  from  the  floccose  aerial  mycelium  growing  upon  gelatin;  conidial  fructifi- 
cations mostly  once  or  twice  alternately  branched,  branches  convergent  or  divergent, 
each  bearing  a  verticil  of  branchlets  crowned  by  verticils  of  conidiiferous  cells  with 
chains  of  conidia,  the  whole  60-240/i,  usually  100-200,«,  in  length;  conidiiferous  cells 
8-10  or  even  13  by  3/x;  conidia  elliptical  or  cylindrical,  then  globose,  3.2-3.5  by  4-4.3/i 
or4/£in  diameter,  adhering  ip  chains  in  fluid  mounts;  grows  luxuriantly  in  fluid  offer- 
ing milk-sugar  as  source  of  carbon,  partially  and  slowly  liquefies  gelatin,  with  alkaline 
reaction  to  litmus;  odor,  very  strong,  "moldy,"  characteristic. 

This  species  was  obtained  from  cheese  sent  from  France  by  Georges 
Roger.  It  was  afterwards  obtained  from  other  French  cheeses,  but 
does  not  seem  to  have  any  economic  importance.  It  is  closely 
related  by  cultural  characters  as  well  as  morphology  to  P.  camem- 
herti,  with  which  it  shares  the  ability  to  grow  normally  in  fluid  offering 
lactose  as  the  source  of  carbon,  but  differs  in  its  short  conidiophores, 
diverse  habit  upon  potato  agar  and  gelatin,  and  its  strong  charac- 

«The  author  is  indebted  to  Prof.  H.  R.  Monteith,  of  the  Connecticut  Agricultural 
College,  for  much  assistaiice  in  preparing  the  Latin  diagnoses. 


PENICILLIUM  BIFORME. 


55 


teristic  smell.  It  is  perhaps  intermediate  in  character  between 
P.  camemberti  and  the  group  of  forms  designated  as  P.  commune  in 
this  paper. 

CULTURAL  DATA. 

Color,  white  to  gray-green,  gray,  or  drab  in  Bome  media,  all  brown  or  drab  when  old; 
reverse  cream;  color  in  media,  none. 

Odor,  very  strong  peculiar  "moldy"  odor,  typical  of  this  species  under  nearly  all 
conditions. 


Fig.  IS.—PenicHlium  biforme:  a,  b,  g,  branching  of  conidial  fructification  from  potato-agar  culture  (x  1,400); 
c,  d,  e,  f,  conidiiferous  cells  and  conidia  (x  1,400);  ft,  j,  k,  sketches  of  conidial  fractifications  on  i>otato 
agar  (X  140);  I,  m,  sketches  of  conidial  fructifications  on  sugar  gelatin  (X  140);  o,  r,  germination  of 

.    conidia  (X  900). 


Fifteen  per  cent  gelatin  in  water,  typical  floccose  colony;  liquefaction,  partial  in 
old  acidified  cultures,  none  in  15  days;  litmus  reaction,  strongly  alkaline  (in  nearly  all 
media) .  Potato  agar  and  bean  agar,  colonies  consisting  only  of  very  short  conidiophores 
green  to  drab  in  color,  little  or  no  floccose  mycelium.  Potato  plugs,  typical.  Cohn's 
solution,  slow,  half  normal  growth,  with  odor. 

SjTithetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  typical  up  to  50  per  cent 
sugar,  with  acid  reaction.     Lactose  3  per  cent,  rich  growth,  typical.     Lactic  acid  0.9 


66  CULTURAL  STUDIES  OP   SPECIES  OF   PENlClLLlUM. 

per  cent,  typical,  alkaline  reaction  when  old,  crystal  drops.  Levulose  3  per  cent, 
typical.  Galactose  3  per  cent,  slow  development,  typical.  Glycerin  3  per  cent, 
typical.  Potato  starch  3  per  cent,  typical  colony,  crystal  transpiration  drops.  Butter- 
fat,  typical  colony,  mycelial  mass  tinged  violet. 

Milk,  rich  growth;  curdling  (0.25  per  cent  calcium  chlorid  added),  in  8  days;  diges- 
tion, rather  slow;  color  in  milk,  none. 

At  37°  C,  no  growth,  slowly  green  when  cooled;  check  at  20  °  C,  typical. 

PENICELLIUM  COMMUNE  n.  sp. 

In  examining  numerous  Petri-dish  cultures  made  for  the  examina- 
tion of  milk  by  the  bacteriologists  of  the  Storrs  Experiment  Station, 
large  numbers  of  colonies  of  Penicillium  have  been  studied.  A  very 
large  percentage  of  these  colonies  have  a  series  of  common  characters 
which  are  constant  enough  to  mark  out  a  species,  or,  perhaps  better,  a 
group  of  races,  between  which  differences  are  either  minute  or  so 
complicated  by  the  occurrence  of  other  races  with  overlapping  char- 
acters as  to  make  their  separation  a  matter  of  considerable  doubt. 
One  of  these  has  been  selected  as  the  basis  of  the  following  diagnosis. 
This  form  is  morphologically  closely  similar  to  P.  expansum  (see 
figs.  1  and  19).  It,  however,  lacks  entirely  the  ability  to  form  core- 
mia  and  fails  to  discolor  the  substrata,  but  grows  well  in  certain  cul- 
ture solutions  which  markedly  restrain  P.  expansum,  of  which  it  might 
possibly  be  regarded  as  a  variety.  From  its  abundance  in  the  situa- 
tions studied  it  has  been  designated  as  P.  commune. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  Solani  tuberosi  aut  phaseoli  cultis, 
viridibus,  demum  brunneolis,  in  eubstrato  late  crescentibus,  zonatis;  marginis  cres- 
centis  parte  aeria  ex  conidiophoris,  centri  atque  ex  hyphis  plus  minusve  floccosis 
composita;  reverse  et  substrato  incolorato;  conidiophoris  plerumque  300/t  raro  usque 
lOO/i  longis;  fructibus  conidicis  100-200/t  longis,  cum  ramis  altcrnatis  et  verticillatis 
confertis,  basidiis  8-9X3^  cum  apicibus  brevibus  acutis,  catenas  conidiorum  longas 
parallelis  gerentibus;  conidiis  primum  cylindricis  vel  ellipticis,  demum  globosia, 
3-4/i  diam.,  5-6/i  incrassatis  germinantibus,  levibua,  viridibus,  in  catenis  manentibua 
Bubmersis;  coloniis  in  gelatina  in  parte  lente  liquefacientibus;  odore  mucidis. 

Habitat  in  lacte,  caseo,  etc.,  Storrs,  Conn. 

Colonies  in  gelatin  or  in  potato  or  bean  agar,  dull  green,  becoming  brown  when  old, 
broadly  spreading,  zonate,  with  broad  white  growing  margin  composed  only  of  conid- 
iophores,  in  the  older  parts  becoming  floccose  masses  of  interwoven  hyphae;  reverse 
of  colony  and  substratum  never  colored ;  conidiophores  commonly  300/«  or  less  in  length, 
sometimes  up  to  700/t;  conidial  fructifications  commonly  100-200jk  in  length,  compact 
at  base  and  broadening  above,  yariously  branching  with  branches  appressed,  and  verti- 
cils of  conidiiferous  cells  8-9  by  3/i,  abruptly/  narrowed  to  produce  conidia;  conidia 
cylindrical  to  elliptical  and  finally  globose  3-4;t,  becoming  4-Ofi  or  larger  in  germinat- 
ing, smooth,  green,  persisting  in  chains  in  fluid  mounts;  colonies  liquefy  gelatin  slowly 
or  partially,  softening  rather  than  producing  clear  liquid,  alkaline  in  media  without 
sugar  but  acid  with  either  cane  sugar  or  lactose,  having  a  strong  "  moldy  "  odor. 

Habitat,  common  in  food,  dairy  products,  etc.,  Storra,  Conn. 


PENICILLIUM   COMMUNE. 


57 


CULTURAL   DATA. 

Color  green,  becoming  brown  when  old;  reverse  cream  or  white;  color  in  media, 
none. 

Odor,  strong,  "moldy." 

Fifteen  per  rent  gelatin  in  water,  good  growth;  liquefaction,  none  in  15  days,  slow  or 
partial  in  older  cultures;  litmus  reaction,  slowly  alkaline.  Potato  agar  and  bean  agar, 
characteristic.  Potato  plugs,  characteristic.  Raulin's  fluid,  characteristic,  with 
odor  "moldy."    Cohn'a  solution,  small  colonies,  trace  of  pink  below. 


\Mv.  kV 


JIG.  19. 


-PeniciUium  commune:  a,  b,  c,  d,  e,  conidial  fructification,  branching,  and  production  of  conldia 
(X  900);  /,  g,  h,j,  k,  I,  sketches  of  fructiflcations  in  various  stages  (X  140). 


Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  typical  culture,  transpired 
drops  crystal.  Lactose  3  per  cent,  typical  culture.  Lactic  acid  0.9  per  cent,  good 
growth.  Levulose  3  per  cent,  good  culture,  strong  odor.  Galactose  3  per  cent,  good 
cultiu-e,  strong  odor,  alkaline  reaction.  Glycerin  3  per  cent,  typical,  strong  odor. 
Butter  fat,  typical  growth. 

Milk,  good  typical  colony;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  1  week; 
digestion,  slow;  color  in  milk,  none. 

At  37°  C,  no  growth,  grew  upon  cooling;  check  at  20°  C,  rich  growth. 


58  CULTURAL  STUtHES  OP  SPECIES  OE  PENIClLLIUM. 

PENICILLIUM  No.  22. 

Colonies  in  gelatin  or  agar  gray-green  or  glaucous  persistently,  or  becoming  gray, 
not  brown,  otherwise  appearing  as  P.  commune;  conidial  fructifications  more  loosely 
branching,  with  branches  divergent;  conidia  larger  and  lighter  color;  odor,  none  or 
indefinite;  reactions  as  in  P.  commune. 

Habitat:  Isolated  several  times  from  domestic  soft  cheeses  made  in  the  State  of  New 
York,  1904-5;  found  associated  with  P.  coTw^mfterh';  in  appearance  and  color  resembling 
latter  species,  but  in  structure  of  colony,  measurements  of  conidiophores,  etc.,  resem- 
bling P.  commune.  In  pure  culture  this  form  has  maintained  its  identity  clearly  for 
four  years. 

CULTURAL  DATA. 

Color  gray-green;  reverse  colorless;  color  in  media,  none. 

Odor,  none  or  very  slight. 

Fifteen  per  cent  gelatin  in  water,  good  growth;  liquefaction,  none  in  15  days,  partial 
in  2  to  3  weeks  or  more ;  litmus  reaction,  blue.  Potato  agar  and  bean  agar,  good  typical 
colonies,  with  alkaline  reaction  to  litmus.  Potato  plugs,  good  growth,  typical  gray- 
green  colony  with  crystal  drops  of  exuded  water.  Cohn's  solution,  medium  growth, 
trace  of  pink  in  media. 

Synthetic  fluid  (Box's),  carbon  supplied  as:  Cane  sugar,  good  growth  up  to  concen- 
tration of  60  per  cent,  acid  reaction.  Lactose  3-10  per  cent,  typical.  Levulose3  per 
cent,  good  growth.  Galactose  3  per  cent,  good  growth,  typical.  Glycerin  3  per  cent, 
good  growth.     Butter  fat,  rich  growth,  typical. 

Milk,  typical  colonies;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  8  days; 
digestion,  rather  slow;   color  in  milk,  none. 

At  37°  C,  no  growth,  grew  when  coeled;  check  at  20°  C,  typical. 

PENICILLIUM  CHRYSOGENUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  Solani  tuberosi  aut  phaseoli  cultie, 
griseo-viridibus,  demum  brunneolis,  in  substrato  late  crescentibus,  margine  sterili 
lata  juvenilibus  parte  aeria  ex  conidiophoris  et  caespitibus  sparsis  hyphorum  ad- 
Bcendentium  composita;  reverse  incolorato;  conidiophoris  plerumque  singulatim 
usque  300X4/t  orientibus,  raro  brevibus  ex  hyphis  assurgentibus  ramosis;  fructibus 
conidicis  100-200  longis  cum  1-2  ramis  alternatis  et  divei^entibus  ramulos  1-2  verti- 
cillatos  gerentibus;  basidiis  8X2. 5/i  verticillatis  ex  apicibus  ramulorum,  catenas 
divergentes  conidiorum  gerentibus;  conidiis  primum  cylindricisvel  ellipticis,  demum 
globosis,  3^  diam.,  pallide  glaucis,  magnis  vacuolis;  coloniis  gelatinam  liquefac- 
ientibus,  alkalinis  lacmo;    lactem,  panem,  gelatinam,  auream  colorantibus. 

Habitat,  in  caseo,  pane,  etc.,  commune. 

Cultivated  in  gelatin,  or  bean  or  potato  agar,  gray-green,  becoming  brownish  when 
old;  aerial  portion  consisting  of  conidiophores  with  some  tufts  of  trailing  aerial  hyphse; 
broadly  spreading  in  the  substratum  with  a  wide  sterile  margin  when  young.  Reverse 
of  colony  not  discolored.  Conidiophores  mostly  arising  separately,  up  to  300  by  4/i, 
partly  arising  as  short  branches  of  aerial  hyphae;  conidial  fructifications  100-200/i  in 
length,  with  1-2  alternate  divergent  branches,  bearing  alternate,  verticillate  or  twice 
verticillate  branchlets.  Conidiiferous cells  8  by  2. 5/t  verticillate  at  the  ends  of  branch- 
lets  bearing  divergent  chains  of  conidia.  Conidia  cylindrical  or  elliptical  at  first, 
then  globose,  3-4/t  in  diameter,  pale  green,  with  large  vacuoles.  Colonies  liquefy 
gelatin,  with  alkaline  reaction  to  litmus  and  produce  in  milk,  bread,  gelatin,  and 
other  substances  a  golden  yellow  celor  (from  which  the  name). 

Habitat,  in  bread,  cheese,  etc.,  apparently  common  and  appearing  in  several 
varieties  which  differ  in  the  intensity  of  color  production,  in  appearance  in  certain 
cultures,  but  which  are  so  far  scarcely  distinguishable  by  structural  characters. 


PENICILLIUM   CHRYSOGENUM. 


59 


The  culture  here  named  P.  chrysogenum  has  been  kept  under 
observation  for  more  than  four  years  without  change.  In  this  time 
several  forms  have  been  collected  or  sent  to  this  laboratory  which 
also  produce  the  golden  color  in  the  digestion  of  milk  suggestive  of 
the  name  proposed.  The  cultural  characters  of  these  races  differ 
in  some  degree  in  several  cases,  in  others  substantial  identity  has 
been  observed.  In  reporting  cultural  data  three  numbers  are 
included  for  comparison  with  this,  viz,  Nos.  25,  35,  and  44.  These 
agree  in  the  following  characters:  Spreading  habit,  surface  mostly 


a^ 


/ 


Fig.  20. — Penicillium  chrysogenum:  a,  b,  c,  d,  e,  branching  of  conidial  fructification  from  gelatin  plates 
(X  900);  /,  g,  h,j,  /.m,  sketches  of  conidial  fructifications  from  potato-agar  plates  (X  140);  n,o,  germination 
ofconidia(X900). 

of  conidiophores  averaging  about  300/t  in  length,  nearly  the  same 
shade  of  green  in  color,  similar  morphology  of  conidial  fructification, 
digestion  of  milk,  gelatin,  etc.,  with  the  production  of  yellow  or 
golden  color,  liquefaction  of  gelatin  progressive  but  slower  than  the 
expansion  of  the  colony  in  the  substratum  so  that  a  growin;^  border 
of  submerged  vegetative  hyphae  extends  into  hard  gelatin  for  con- 
siderable time,  in  contrast  to  forms  in  which  the  colony  becomes  a 
floating  "island"  in  a  pool  of  fluid  within  the  first  week.  In  spite 
of  these  common  characters  colonies  of  these  four  races  often  show 


60  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

^TOiit  apparent  difTeronces  in  parallel  culture  in  some  specialized 
media.  It  seems  preferable  to  let  this  name  stand  in  a  broad  enough 
sense  to  include  the  forms  having  the  common  characters  above 
noted  than  to  attempt  any  narrower  delimitation  of  this  group  of 
forms  at  this  time. 

Serial  No.  57. — Another  form  which  may  be  included  temporarily 
with  this  group  does  not  produce  a  yellow  color  in  the  substratum 
at  all,  but  produces  mycelium  of  orange  color  as  seen  from  below. 
The  surface  growth  is  clear  green  without  definite  differences  from 
the  chrysogenum  group,  but  the  orange  color  in  reverse  remains  as 
a  constant  difference  from  that  group,  unattended  by  any  coloration 
of  the  medium. 

CULTURAL    DATA. 
(Of.  Nos.  25,  35,  and  44  in  tables.) 

Color  gray -green,  green,  to  brown  when  old;  reverse  colorless,  or  slight  yellowish. 

Color  in  media  golden  yellow  in  certain  media,  no  color  in  others. 

Odor,  none. 

Fifteen  per  cent  gelatm  in  water,  good  growth,  yellow  color  in  gelatin;  lique- 
faction rapid — 6-10  days,  more  or  less  rapid  and  complete  in  all  gelatin  media  used; 
litmus  reaction,  alkaline.  Potato  agar  and  bean  agar,  good  colonies,  but  no  yellow 
color.  Potato  plugs,  typical,  potato  becoming  yellow.  Raulin's  fluid,  typical  with 
yellow  transpired  drops,  but  no  yellow  in  fluid.  Cohn's  solution,  growth,  but  not 
normal. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  growth  up  to  30  per 
cent,  reaction  acid,  fluid  colorless.  Lactose  3  per  cent,  fair  growth,  not  heavy,  fluid 
yellow,  with  acid  reaction.  Lactic  acid  0.9  per  cent,  some  growth.  Galactose  3 
per  cent,  good  growth.  Glycerin  3  per  cent,  colony  white  to  green,  no  yellow  color 
in  fluid.  Potato  starch  3  per  cent,  good  growth,  yellow  drops  above,  yellow  in  fluid. 
Butter  fat,  rich  growth,  with  fluid  golden  yellow. 

Milk,  rich  growth ;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  1  week ;  digestion 
rapid;  color  yellow. 

PENICILLIUM  RUGULOSUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  phaseoli  cultis,  flavo-viridibus,  dein 
viridibus,  demum  atro-viridibus,  late  crescentibus  in  agaro;  parte  aeria  ex  conidioph- 
oris  creberrimis  et  hyphisaereis  et  paucis  composita;  reverse  luteo  et  in  parte  aurantiaco 
imprimis  in  tubere  Solani;  conidiophoris  100-200 X 2. 5-3/t,  singulatim  vel  ex  hyphis 
aereis  prope  substratum  orientibus;  fructibus  conidicis  100-150/t  longis  (in  saccharo 
nulto longioribus)  ex  ramis  10-15x2. 5/z,  compacte  verticillatis,  verticillos  basidiorum, 
vel  ramulorum,  vel  ramulorum  et  basidiorum  eodem  verticillo  gerentibuf;  basidiis 
9-12  X2/(,  acuminatis,  catenas  longas  et  divergentes  conidiorum  gerentibus;  conidiis 
3.4-3.8X-2.5-3j((,  ellipticis,  viridibus,  uno  apice  incrassato,  verruculosis  maturis,  in 
catenis  manentibus  submersis,  5/i  diam.  incrassatis  germinantibus;  coloniis  non  (vel 
solum  in  parte  et  lente)  gelatinam  liquefacientibus. 

Commune  in  culturis,  Storrs,  Conn. 

Cultivated  in  gelatin  or  bean  agar,  yellowish  green,  then  green,  at  length  dark  green; 
surface  growth  of  densely  crowded  conidiophores  with  few  aerial  hyphae  interspersed 
at  their  bases;  reverse  of  colonies  yellow  to  orange  in  spots,  especially  upon  potato 
or  upon  sugar  media;  substratum  not  or  slightly  yellowed;  conidiophores  100-200 
by  2.5-3  ju,  arising  separately  or  branching  from  aerial  hyphae  just  above  the  substratum ; 
conidial  fructifications  100-150;t  in  length,  consisting  of  appressed,  verticillate  branches 


PENICILLIUM   RUGULOSUM. 


61 


10-15  by  2.5/1,  bearing  verticils  of  conidiiferous  cells,  of  branchlets,  or  of  conidiiferous 
cells  and  branchlets  together;  conidiiferous  cells  9-12  by  2;t,  acuminate,  bearing  long 
divergent  chains  of  conidia;  conidia  3.4-3.8  by  2.5-3/t,  elliptical,  green,  mostly  with 
swelling  at  one  end,  verruculose  when  ripe,  swelling  to  5/i  and  germinating  by  one 
or  two  tubes;  colonies  do  not  or  only  partially  liquefy  gelatin. 

Common  in  cultures,  Storrs,  Conn.  Characterized  by  its  verrucose  spores  and  the 
brilliant  color  of  the  mycelium  viewed  from  below. 

CULTURAL   DATA. 

Color  green  or  yellowish  green;  reverse  yellow  to  orange  or  reddish  orange;  color 
in  media,  none. 

Fifteen  per  cent  gelatin  in  water,  slow  development;  liquefaction,  none  or  slight 
in  old  colonies;  litmus  reaction  neutral.  Potato  agar,  good  growth,  alkaline.  Bean 
agar  +  5  per  cent  cane  sugar,  rich  heavy  growth  producing  conidia  in  heavy  dark- 
green  masses,  easily  broken  off  by  shaking  the  tube.  Potato  plugs,  green,  reverse 
bright  orange. 


FiQ.  21. — Penicillium  rugulomm:  a,  h,  branching  of  conidiophore  (X  1,600);  c,d,  e,  conidiiferous  cells  (basidia) 
and  conidia  (X  1,600);  /,  fully  ripeconidium,  showing  delicate  roughening  of  walls  (X  l,e00);  g,  h,j,  swell- 
ing and  germination  of  conidia  (X  1,600);  /,  m,  diagrams  of  conidial  fructifications  (X  260). 

Synthetic  fluid  (Dox's)^  carbon  supplied  as:  Cane  sugar,  growth  in  concentration 
up  to  60  per  cent,  acid.  Lactose,  10  per  cent,  slight  growth.  Levulose,  fair  growth, 
acid  reaction ;  galactose,  good  growth;  glycerin,  slow  growth;  butterfat,  slowly  typical 
colony,  yellow  below;  fluid,  slightly  yellowish. 

Milk,  growth  typical,  reverse  yellow  to  orange  or  reddish  orange;  curdling  (0.25 
per  cent  calcium  chlorid  added)  in  9  days;  digestion  slow  or  slight;  color,  none  in 
milk. 

At  37°  C,  no  growth,  grew  when  cooled;  check  at  20°  C,  typical. 

PENICILLIUM  CITRINUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  Solani  tuberosi  aut  phaseoli  cultis, 
aeruginoso-viridibus,  demum  fuligineis;  fructibus  viridibus  usque  ad  marginem  gestis, 
i.e.  margine  sterile  angustissima;  coloniis  in  gelatina  rotundis,  parvis,  cito  liquefa- 


62 


CULTURAL  STUDIES  OF  SPECIES  OF  PENICILLIUM. 


centibus;  in  agaro  latioribus;  parte  aeria  ex  conidiophoria  et  fructibus  conidiciscreber- 
rimis  composita,  interdum  caespitibus  paucis  hyphorum  adscendentum  in  medio; 
reverso  incolorato;  conidiophoria  (sine  ramia)  non  longioribus  150/i,  singulatira  orienti- 
bu8,  aut  paucia  ex  hyphia  adacendentibua  ramoaia;  fruotibua  conidicia  3-5  ramorum 
16-30X3^,  apice  5fi  incrassatorum,  in  verticillo,  basidia  in  verticillia  compactia 
gerentum;  utroque  verticillo  catenia  conidiorum  in  columno  conipacto  50-150/x 
longos  adhaerentibus;  baaidiia  6-7X2-3/i;  conidiia  globosia,  2.4-3^  raro  3.5/x  diam., 
aeruginoao-glaucis,  granulatia  intus,  in  catenia  manentibus  aubmeraia. 

Coloniis,  eaccharo  commixto,  substrata  citrina  in  colore  efficientibus  (ubi  nomen). 

Habitat,  in  caseo,  pane,  etc.,  commune  (?). 

Coloniea  grown  upon  gelatin  and  potato  or  bean  agar  blue-green  when  young, 
becoming  dark  brown  when  old,  with  colored  fruit  borne  almost  to  the  very  margin 


FlO.  22.—Pe7ucillium  citrinum:  a,  b,  e,  /,  g,  branching  of  conidial  fructification,  showing  number  of 
branches  in  each  verticil  and  enlargement  of  ends  of  branches  (o  X  900,  others  X  1,600);  c,  d,  n,  conidiif- 
erous  cells  and  the  formation  of  conidia  (X  1,600);  h,  j,  k,  I,  m,  sketches  of  conidial  fructifications 
(X  140);  o,  s,  t,  germination  of  spores  (X  900). 


of  the  colony,  so  that  the  white  border  of  submerged  mycelium  and  uncolored  fruit 
is  very  narrow;  restricted  in  growth  to  a  few  millimeters  in  diameter  upon  gelatin, 
but  becoming  much  larger  upon  agar;  aerial  part  of  colony  consisting  of  densely 
standing  conidiophores  and  conidia  except  in  the  center,  where  there  arise  a  few  tufts 
of  trailing  aerial  hyphae.  Reverse  of  colony  itself  colorless  or  only  yellowish.  Conidio- 
phores arising  separately,  rarely  longer  than  ISOju,  branching  acropetally  from  sub- 
merged hyphse  radiating  from  the  center  of  the  colony,  or  branched  from  the  hyphae 
of  the  central  aerial  tuft.  Conidial  fructification  a  verticil  2  to  5  branches  16-30  by  3// 
enlarged  at  apex  to  5/u,  each  producing  a  compact  verticil  of  conidiiferous  cells  bearing 
chains  of  conidia  massed  together  into  columns  50-150/1  in  length  (usually  80-100/i). 


PENICILLIUM   NO.  37.  63 

The  whole  fructification  appears  in  this  way  double,  triple,  or  quadruple  or  even 
more  complex  by  a  secondary  verticil  from  the  central  branch.  Conidiiferous  cells 
6-7  by  2-3(1.  Conidia  globose  when  ripe,  2.4-3/1  (even  3.5;t  diam.  in  cane-sugar  cul- 
tures) in  diameter,  bluish-green,  slightly  granular  in  contents,  adhering  in  chains 
in  fluid  mounts,  losing  vitality  rapidly  with  change  of  color  in  old  colonies.  Colonies 
liquefy  gelatin  rapidly,  so  that  they  lie  in  pools  of  liquid  within  a  week.  Litmus 
reaction  in  plain  gelatin,  strongly  alkaline.  Produces  a  lemon-yellow  color  soluble 
in  alcohol  in  media  containing  sugars,  milk,  gelatin,  bread,  and  potato. 

Found  in  cultures  from  milk  and  cheese,  probably  cosmopolitan. 

Could  this  be  P.  citreo-nigrum  Dierckx? 

CULTURAL   DATA. 

Color  bluish  green,  becoming  dark  bro^n  when  old  if  exposed  to  light;  reverse 
colorless  or  yellowish;  color  in  media  lemon-yellow  when  cane  sugar  or  gelatin  or 
p>eptone  is  present,  none  in  some  media. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  characteristic  growth,  becoming  white  by  sec- 
ondary sterile  mycelium;  liquefaction  rapid,  colonies  floating  in  5-6  days;  litmus 
reaction  alkaline.  Potato  agar  and  bean  agar,  colonies  spreading  more  than  upon 
gelatin,  agar  not  colored  or  slightly  colored.  Potato  plugs,  typical  growth,  potato 
colored  yellow,  with  yellow  drops  transpired;  Raulin's  fluid,  typical  growth,  very 
slight  yellow  color.     Cohn's  solution,  good  growth,  no  yellow  color. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  growth  with  yellow 
color  up  to  30  per  cent,  with  acid  reaction.  Lactose  3  per  cent,  slow  poor  growth 
giving  a  violet  tinge  instead  of  yellow  color.  Lactic  acid  0.9  per  cent,  slight  gro\vth. 
Levulose  3  per  cent,  small  characteristic  growth.  Galactose  3  per  cent,  good  growth, 
reaction  neutral  or  slightly  acid.  Glycerin  3  per  cent,  v«ry  small  colonies,  no  color 
in  medium.  Alcohol,  5  per  cent,  good  growth,  no  yellow.  Potato  starch  3  per  cent, 
good  growth,  slight  if  any  yellow.  Tartaric  acid,  slow  colorless  colony.  Biitterfat, 
rich  growth,  lemon-yellow  fluid. 

Milk,  rapid  growth;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  10  days; 
digestion  rapid;  coloration  pale  yellowish. 

At  37°  C,  slow  growth,  white  colony,  no  color  in  medium;  check  at  20°  C,  rich 
growth,  green,  yellow  in  medium,  in  bean  agar  with  cane  sugar. 

PENICILLIUM  No.  37. 

(Var.  of  P.  citrinumt    Or  allied  to  P.  cUrinumt) 

Colonies  in  media  without  sugar,  green,  gray -green,  or  gray;  with  sugar  persistently 
green;  surface  velvety  strict,  composed  of  short  crowded  conidiophores  up  to  lOO/i  in 
length,  branching  from  closely  woven  mycelium  partly  submerged,  partly  aerial, 
with  margin  narrow,  not  widely  spreading  in  the  substratum;  reverse  of  colony  and 
substratum  not  colored  or  creamy;  conidial  fructifications  sometimes  a  single  verticil 
of  conidiiferous  cells,  sometimes  2  to  4  verticillate  branches;  chains  of  conidia  from 
each  verticil  forming  a  column  up  to  500-600/1  in  length  in  sugar  media;  branches 
of  fructification  13-14  by  2-2.5/1  enlarged  at  apex;  conidiiferous  cells  8-10  by  2+/t 
abruptly  narrowed  into  sterigmata,  usually  6-10  in  each  verticil;  conidia  broadly 
elliptical  to  globose,  2.5-3/t  at  first  becoming  4-5/i  before  germinating,  thin-walled, 
smooth,  pale  yellowish  green,  germinating  by  a  single  tube;  colonies  liquefy  gelatin 
rapidly  (6  to  7  days),  with  strong  alkaline  reaction  to  litmus. 

Received  from  Prof.  P.  H.  Rolfs,  Miami,  Fla.,  in  culture  upon  bean  stems,  1905. 

Allied  to  P.  citrinum  by  morphology  and  culture  reactions,  but  differing  in  lacking 
the  power  to  color  media  yellow  and  in  its  greater  dependence  for  typical  growth 
upon  the  presence  of  cane  sugar. 


64 


CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 


CULTURAL   DATA. 

Color  light  blue-green,  olive,  or  gray  in  various  media;  reverse,  white  or  cr^am; 
color  in  media,  none  or  slightly  yellowish. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  small  olive-green  colonies;  liquefaction,  rapid — 
6  to  7  days;  litmus  reaction,  alkaline.  Potato  agar  and  bean  agar,  typical.  Potato 
plugs,  typical.  Raulin's  fluid,  typical.  Cohn's  solution,  slow  and  weak-growing 
colony. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  growth  even  up 
to  60  per  cent,  acid  reaction.  Lactose,  3  per  cent,  slow  abnormal  growth.  Lactic 
acid,  0.9  per  cent,  small  but  characteristic  colony.     Levulose,  3  per  cent,  small  colo- 


FiG.  23.— Penicillium  No.  37:  a,  typical  branched  fructification,  two  verticils  ofconidiiferousceUs(Xl,600); 
b,c,  conidiiferous  cell  and  conidia  (X  1,600);  d,  e,  sketches  of  conidial  fructifications  from  potato-agar 
culture  (X  HO);  /,  g,  sketches  from  an  old  culture  on  3  per  cent  cane-sugar  agar,  showing  simple  (j)  and 
branched  (/)  forms  (X  140);  ft,  j,  fc,  branching  of  conidiophore,  swollen  ends  of  branches  (X  1,000);  I,  m, 
germination  of  conidia  (X  1,600). 

nies.  Galactose,  3  per  cent,  typical.  Glycerin,  3  per  cent,  small  growth.  Potato 
starch,  typical.     Butterfat,  slow  growth,  deep  heavy  green  colony. 

Milk,  curdling  (0.25  per  cent  calcium  chlorid  added),  rapid ;o  digestion  rapid  and 
complete;  color,  none. 

At  37°  C,  killed;  check  at  20°  C,  grew  well. 


PENICILLIUM  No.  12. 

This  form  differs  "from  P.  citrinum  in  producing  no  coloration  of  the  medium  and 
in  producing  conidial  fructifications  in  which  the  chains  of  conidia  are  more  or  less 
divergent  instead  of  aggregated  into  columns.     In  culture  there  is  general  corre- 

aThe  time  of  curdling  is  almost  impossible  to  determine  in  cases  where  digestion 
begins  quickly  and  progresses  rapidly. 


•      PENICILLIUM  ATEAMENTOSUM.  65 

spondence  in  reactions,  without  identity;  this  form  appears  to  be  much  more 
dependent  upon  cane  sugar  for  the  production  of  typical  color  of  the  conidia  and 
growth  than  is  P.  citrinum.  There  is  also  a  greater  tendency  to  the  production 
of  a  layer  of  mycelial  hyphae  just  above  the  surface  of  the  substratum,  from  which  the 
conidiophores  arise  as  aerial  branches. 

This  form  was  received  from  Prof.  C.  E.  Marshall,  Agricultiual  College,  Michigan 
under  the  name  of  P.  glaucum. 

CULTURAL   DATA. 

Color  pale  blue-green;  reverse  of  colony  cream,  not  colored;  color  in  media,  none. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  rather  small  pale  blue  colonies,  rapidly  becoming 
white  by  secondary  sterile  growth  of  hyphse;  liquefaction,  rapid — 6  days;  litmus 
reaction,  alkaline.  Potato  agar,  as  in  gelatin.  Potato  plugs,  very  poor  growth, 
grayish  or  yellowish  green.  Raulin's  fluid,  slow  but  typical  colonies,  deUcate  blue; 
Cohn's  solution,  slow  and  restricted  growth. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  grows  well  in  concentra- 
tions up  to  60  per  cent.  Lactose  3  per  cent,  small  colonies  lacking  nourishment. 
Lactic  acid  0.9  per  cent,  small  colonies  floating  in  fluid.  Levulose  3  per  cent,  good 
growth,  alkaline  reaction.  Galactose  3  per  cent,  good  growth,  alkaline  reaction. 
Glycerin  3  per  cent,  slow-growing  colonies,  becoming  gray-brown  when  old .  Potato 
starch,  good  colonies.     Butterfat,  slow  and  ill-nourished  growth. 

Milk,  rapid  growth;  curdling  (0.25  per  cent  calcium  chiorid  added)  in  7  days;  diges- 
tion rapid  and  very  complete;  color  in  milk,  none. 

At  37°  C,  killed  in  6  days;  at  20°  C,  good  growth. 

PENICILLIUM  ATRAMENTOSUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  Solani  tuberosi  aut  phaseoli  cultis, 
viridibus,  parte  aeria  plerumque  ex  conidiophoris  singulatim  orientibus,  medio  cum 
hyphis  aereis  interspersis,  margine  albo  ex  hyphis  fertilibus  angusta;  reverso  incolorato 
vel  parum  ochraceo;  substrato  aut  incolorato  aut  in  substratis  saccharinis  et  in  lacte 
atrobrunneo  tarde  fere  atro;  conidiophoris  240-300  usque  400/z  longis;  fructibus  conidi- 
cis  100  usque  200/i  longis,  ramis  1-2  verticillatis  2-4  insequaliter  longis  in  verticillo  in 
apice  incrassatis;  basidiis  8-10/t  longis,  parallelis  in  verticillo;  catenis  conidiorum 
eodem  verticillo  in  columno  compactis;  conidiis  ellipticis,  3.5-4' (usque  4.8)  X2.5-3 
usque  3.5/i,  Isevibus,  viridibus,  6-7/i  incrassatis  et  uno  tubo  germinantibus;  coloniis 
gelatinam  cito  liquefacientibus,  alkalinis  lacmo;  odore  in  lacte  proprio,  in  substratis 
aliis  nuUo. 

Ex  caseo  cultum,  Storrs,  Conn.,  1905. 

Afiine  P.  citrino. 

Colonies  upon  gelatin  or  upon  potato  or  bean  agar  bright  green,  aerial  part  mostly 
of  simple  conidiophores,  mixed  in  older  parts  with  branching  aerial  hyphae  but  nar- 
rowly spreading  at  the  margin  by  new  conidiophores  only.  Reverse  of  colonies  shows 
a  slight  production  of  yellow  (ochraceous)  color.  Conidiophores  240-400/u,  averaging 
■about  300//  in  length.  Conidial  fructification  up  to  200/i  in  length,  usually  lOO/x  or 
less,  verticillately  or  twice  verticillately  branched;  branches  2-4  in  a  verticil  di- 
vergent, unequal  in  length,  swollen  at  ends,  bearing  conidiiferous  cells.  The  conidial 
chains  from  each  verticil  form  a  dense  column,  which  diverges  more  or  less  from  the 
other  columns  when  old .  Conidiiferous  cells  8-I0/<  in  length ,  closely  parallel .  Conidia 
elliptical,  varying  from  3.5-4/t  by  2.5-3//  on  agar,  somewhat  larger  in  gelatin  cultures, 
up  to  4.8  by  3.5/i,  smooth,  homogeneous  green  with  a  slight  yellowish  shade  when  seen 
in  mass,  swelling  to  6-7//  in  diameter  and  germinating  by  a  single  tube.  Mycelial 
cells  5-7//  in  diameter  and  up  to  30//  or  more  in  length.  Colonies  liquefy  sugar-gelatin 
8108— Bull.  118—10 5 


66 


CULTURAL  STUDIES   OF   SPECIES  OF   PENICILLIUM. 


rapidly,  give  an  alkaline  reaction  to  litmus,  digest  milk,  and  color  potato  agar  con- 
taining high  p>ercentage  of  sugar  a  deep  black. 

Found  upon  Camembert  cheese  imported  from  France.  Closely  related  morpho- 
logically to  P.  cilrinum,  from  which  it  is  separated  by  the  longer  conidiophores  and 
larger  spores  as  well  aa  the  black  discoloration  of  sugar  media. 

CULTURAL  DATA. 

Color  deep  (blue)  green  to  brown  when  old;  reverse  uncolored,  or  brown  in  some 
media;  color  in  media,  none,  or  brownish  to  almost  black. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  deep  green,  brown  when  old,  rich  growth;  lique- 
faction, 7  days,  varies  from  6  to  12  days  in  other  gelatin  media;  litmus  reaction,  alka- 
line. Potato  agar  and  bean  agar,  typical,  no  color  below.  Potato  plugs,  dark  green, 
potato  blackened.  Raulin's  fluid,  rather  weak  growth.  Cohn's  solution,  germinated 
only. 


Fig.  2i.—Penicilltum  atramerUosum:  a,  6,  c,  d,  branching  of  conidial  fructification  showing  unequal 
length  of  branches,  swollen  ends  (X  900);  e,  f,  conidiiferous  cell  and  chain  of  conidia(X  900);  g,  h,  j, 
sketches  of  conidial  fructifications  (X  HO);  i,  conidia  (X  1,000);  m,  n,  o,  t,  germination  of  conidia 
(X900). 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  growth  up  to  50  per 
cent,  with  acid  reaction  in  50  per  cent  solution.  Lactose  3  per  cent,  small  and  slow 
growth.  Lactic  acid  0.9  per  cent,  no  growth.  liCvulose  3  per  cent,  slowly  typical. 
Galactose  3  per  cent,  typical,  with  alkaline  reaction.  Glycerin  3  per  cent,  germina- 
tion. Potato  starch  3  per  cent,  good  growth,  no  color  in  fluid  or  reverse  of  colony. 
Butterfat,  typical,  green  colonies  with  reverse  brown,  and  fluid  uncolored. 

Milk,  curdling  (0.25  per  cent  calcium  chlorid  added)  in  9  days;  digestion,  rapid, 
fairly  complete;  color,  brownish  to  aimost  black. 

At  37°  C,  no  growth,  grew  when  cooled;  check  at  20°  C,  typical. 

PENICILLIUM  No.  24. 

(Related  ta  P.  atramerUosum?) 

Colonies  upon  gelatin  and  potato  or  bean  agar  blue-green,  becoming  brown  rapidly 
when  old,  or  smoky  with  very  dense  velvety  surface  consisting  of  conidiophores  aris- 
ing in  the  substratum  or  just  above  its  surface,  with  a  very  abrupt  narrow  white 


PENICILLTUM   NO.   24. 


67 


mai^in  of  unripened  fruit  and  submerged  mycelium  during  the  growing  period. 
Reverseof  colony  and  medium  colorless  under  all  conditions  studied.  Conidiophores 
from  100  to  400/i,  averaging  about  250//,  in  length,  either  arising  separately  or  as  lateral 
branches  of  hyphse  just  above  the  substratum.  Conidial  fructification  up  to  200/t  in 
length  produced  by  various  branching  from  the  conidiophores  in  which  each  primary 
branch  is  often  divergent  to  produce  separate  mass  of  conidia.  Conidiiferous  cells  7-10 
by  3/(.  Conidia  globose  3.3-4/t,  homogeneous  blue-green,  smooth,  seeming  to  lose 
vitality  rapidly  under  laboratory  conditions.  Colonies  liquefy  gelatin  in  7  to  12  days 
so  that  they  lie  in  pools  of  liquid.     Litmus  reaction  strongly  alkaline. 

Found  in  the  cultures  from  Camembert  cheese  in  the  laboratories  at  Storrs,  Conn. 
Differs  from  the  preceding  and  from  P.  citrinum  by  its  longer  conidiophores,  the 
alternate  branching  of  its  fructification,  the  size  of  its  spores,  and  by  failure  to  color 
the  substrata.  The  relative  value  of  ellipticity  of  conidia  as  a  diagnostic  character 
appears  to  be  questionable.    This  form  is  therefore  presented  under  cultural  number 


Fig.  2&.—Penicillium  No.  24:  a,  b,  branching  and  arrangement  of  branches  inconidialfructification  (X  140); 
c,  d,  e,  conidiiferous  cells  and  conidia  (X  1,400);  g,  h,j,  sketchesof  form  and  arrangement  of  conidiophores 
(X  140);  tn,  n,  germination  of  conidia  (X  900). 


only,  whereas  the  preceding  has  been  identified  from  accidental  cultures  more  fre- 
quently, and  hence  is  given  name  and  description  as  a  species. 

CULTURAL   DATA. 

Color  deep  green  (blue-green),  becoming  brown  when  old  and  exposed;  reverse 
white  or  cream;  color  in  media,  none. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  characteristic  colony;  liquefaction,  rapid — 
11  days  or  even  less;  litmus  reaction  alkaline.  Potato  plugs,  deep  blue-green,  crystal 
drops.  Raulin's  fluid,  weak  but  characteristic  growth.  Cohn's  solution,  slow  but 
characteristic  development. 

Synthetic  fluid  (Dox's),  carbon  supplied:  Cane  sugar,  good  growth  up  to  30  per 
cent,  acid  reaction.  Lactose,  3  per  cent,  slow  development,  not  typical.  Lactic 
acid  0.9  per  cent,  good  colony.     Levulose  3  per  cent,  small  colonies.    Galactose  3 


68 


CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 


per  cent,  good  growth,  strongly  alkaline.  Glycerin  3  per  cent,  weak  growth.  Potato 
starch,  characteristic  colony. 

Milk,  curdling  (0.25  per  cent  calcium  chlorid  added)  in  8  days;  digestion  medium 
rapid;  color  in  milk,  none  or  ? 

At  37°  C,  no  growth,  grew  when  cooled;  check  at  20°  C,  good  typical. 

PENICILLIUM  STOLONIFERUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  Solaui  tuberosi  cultis,  viridibus  vel 
flavo-viridibus,  demum  griseo-viridibua  vel  griseis  in  agaro  sine  saccharo,  cum  sac- 


Fio.  26.— PentctHmTO  stoloniferum:  a,  b,  c.  e,/.  the  types  of  branching  at  the  tips  of  the  "stolons"  by  which 
this  species  spreads  in  substrata  (6,  c,  e,/.  X  900);  d,  conidial  fructification  ( X  900);  h,j,k,l,  sketches 
ol  conidial  fructifications  of  various  ages,  h  and  j  being  characteristic  shapes  (X  140);  g,  formation  of 
conidia(X  900);  t,  ripe  conidia  showing  minute  granulation  (X  1,600);  m,  n,  germination  of  conidia 
(X  900);  0,  rough  diagram  of  habit. 


charo  viridibus,  floccosis,  in  culturis  juvenalibus  stolonibus  aereiscitius  quam  hyphis 
Bubmersis  crescentibus,  re  verso  incolorato  vel  in  parte  flavo;  conidiophoris  brevibus 
ex  hyphis  adscendentibus  ramosis,  usque  lOO/i  longis,  aut  singulatim  orientibus  (sine 
ramis)  plus  minus  300/x  longis;  fructibus  conidicis  40-80/z  raro  usque  170/i  longis,  ex 
ramls  brevibus  compactis,  et  basidiis  verticillatis,  in  baside  confertissimis,  catenas 
conidiorum  late  divergentes  gerentibus  compositis,  interdura  ramus  infimus  tarn 
divergens  ut  fructus  duplex  videatur;  basidiis  lOX  3;/;  conidiis  ellipticis  vel  paene 
globosis,  2.8-3.4/t  diam.,  pallido  flavo-viridibus  Isevibusque;  coloniis  gelatinam  cito 
liquefacientibus,  alkalinia  lacmo. 


PENICILLIUM  FUNICULOSUM.  69 

Habitat,  in  fungis  putreacentibus,  Boletis,  Polyporis;  Storre,  Conn.;  Paris,  Gallia. 

Cultivated  in  gelatin  or  potato  agar,  green  or  yellowish  green,  becoming  gray-green 
or  gray  when  old  (remaining  green  in  sugar  media),  floccose,  spreading  more  rapidly 
in  young  cultures  by  aerial  stolons  than  by  submerged  hyphte  (i.  e.,  the  submerged 
mycelium  seems  to  arise  from  the  aerial  rather  than  vice  versa);  reverse  of  colony  not 
colored  or  partly  yellow;  conidiophores  arising  as  short  branches  (100/i  or  less  in  length) 
from  aerial  hyphae,  or  arising  separately  300/[t  or  more  in  length  especially  at  the  mar- 
gins of  older  colonies;  conidial  fructification  40-80  more  rarely  up  to  170//  in  length, 
composed  of  short  appressed  branches  and  numerous  conidiiferous  cells  densely 
crowded  at  the  base  bearing  very  loosely  divergent  chains  of  conidia;  sometimes  the 
lowest  branch  diverges  so  that  the  fructification  appears  double;  conidiiferous  cells 
10  by  3ft;  conidia  slightly  elliptical  or  globose,  2.8-3.4/£,  smooth,  yellowish  green  in 
mass,  almost  hyaline  by  transmitted  light;  colonies  liquefy  gelatin  very  rapidly,  with 
a  strong  alkaline  reaction  to  litmus. 

Habitat,  decaying  fungi,  Boleti,  Polypori;  cultures  from  milk  and  ensilage.  Col- 
lected repeatedly  at  Storrs,  Conn.;  once  upon  decaying  Boletus  scaber  at  the  Jardin  des 
Plan  tea  in  Paris,  hence  probably  widely  distributed.  The  stolon-producing  character 
is  so  easily  seen  and  so  characteristic  of  this  species  as  to  seem  adequate  to  distinguish 
it  from  all  other  species  studied.  This  has  been  observed  upon  a  decaying  Boletus 
with  a  hand  lens. 

CULTURAL  DATA. 

Color  white  to  yellowish  green,  deep  green  becoming  yellowish  brown  or  gray  in 
old  cultures;  reverse,  not  colored  (or  slightly  yellow);  color  in  media,  none  or  slight. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  good  growth,  yellowish  green;  liquefaction,  rapid 
in  all  gelatin  media;  litmus  reaction,  strongly  alkaline.  Potato  agar,  good  growth, 
pale  green  to  gray.  Bean  agar,  good  growth,  pale  green  to  gray.  Potato  plugs,  good 
growth,  deep  green,  transpired  drops  brown.  Raulin's  fluid,  slow  but  characteristic 
colonies.     Cohn's  solution,  typical  growth. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  growth  up  to  50  per 
cent,  conidial  areas  persistently  green  (viridis  to  atro-viridis),  acid  reaction.  Lactose 
3  per  cent,  slow  abnormal  colonies,  weak.  Lactic  acid  0.9  per  cent,  small  colonies  be- 
coming alkaline.  Levulose  3  per  cent,  typical  colonies,  alkaline  reaction.  Galactose 
3  per  cent,  typical,  alkaline  reaction.  Glycerin  3  per  cent,  good  growth,  not  heavy. 
Potato  starch  3  per  cent,  typical,  drops  yellow,  fluid  colorless. 

Milk,  typical  colonies;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  1  week; 
digestion,  rapid;  color  in  milk,  little  or  none. 

At  37°  C,  killed;  check  at  20°  C,  good  colony. 

PENICILLIUM  FUNICULOSUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  Solani  tuberosi  aut  phaseoli  cultis, 
atro-viridibus,  late  crescentibus,  floccosis;  parte  aeria  ex  hyphis  decumbentibus, 
ramosis,  csespitosis,  late  intricatis,  et  fasciculatis,  conidiophoros  breves  gerentibus 
interdura  hyphos  secundarias  albas  floccosas  lente  evolvente;  reverso  rubescente 
demun  atro-vinoso;  substrato-aut  lacte-aut  gelatina,  vinoso;  conidiophoris  (sine  ramis) 
20-80  usque  100/z  longis,  plerumque  ex  hyphis  repentibus  vel  fasciculatis,  interdum 
singulatim  orientibus,  fructibus  conidicis  usque  125-160;*  longis,  cum  1,  2  ramis  alter- 
natis,  dein  ramulis  verticillatis,  basidia  in  verticillos  densos  catenis  conidiorum  paral- 
lelis  gerentibus;  basidiis  10-14X2-3/1,  parallelis  in  verticillo,  acuminatis;  conidiis 
primum  cylindricis,  demum  fusiformibusv  el  ellipticis,  3-4X2-3/1,  viridibus;  conidi- 
orum catenis  solventibus  submersis;  coloniis  gelatinam  non  liquefacientibus,  acidis 
lacmo,  siccantibussenescentibusque  interdum  coremiis  paucis  evolventibus. 

In  cultura,  Storrs,  Conn.,  1905;  communicavit  Dr.  £.  A.  Beesey,  Miami,  Fla.,  1908. 


70 


CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 


Cultivated  in  gelatin  or  potato  or  bean  agar,  deep  green,  broadly  spreading,  surface 
closely  floccose  with  procumbent  hyphae,  tufts  and  ropes  of  hyphse  bearing  lateral 
conidiophores;  reverse  becoming  red,  purple,  or  very  dark  purple,  almost  black,  with 
the  whole  mass  of  medium  colored;  conidiphores  short,  20-80  or  100/i,  mostly  perpen- 
dicufar  branches  from  trailing  hyphae,  sometimes  arising  separately  from  the  sub- 
stratum; conidial  fructification  up  to  125  or  IGOfi  in  length,  with  1  or  2  alternate 
appressed  branches  bearing  verticillate  branchlets  and  dense  verticils  of  parallel  coni- 
diiferous  cells  10-14  by  2-3/i;  conidia  at  first  cylindrical,  then  elliptical  or  fusiform, 
3-4  by  2-3/t,  green,  in  chains  which  break  up  completely  in  fluid  mounts;  colonies 
not  liquefying  gelatin  in  2  weeks,  with  acid  reaction  to  litmus. 


Fig.  27.— PenkiUiumfuniculoxum:  a,  b,  c,  d,  t,f,  conidial  fructifications  with  conidiiferous  cells  and  conidia 
(X  900,  except  c,  1,()00);  g,  b,  j,  k,  I,  m,  n,  sketches  of  fructifications,  separate  and  borne  upon  hyphte 
and  ropes  of  hyphae  (X  140);  o,  r,  germination  of  conidia  (X  900). 

Found  in  accidental  culture,  Storrs,  Conn.,  1905;  also  received  from  Dr.  E.  A. 
Bessey,  Miami,  Fla.,  1908.     Easily  recognized  in  culture. 


CULTURAL  DATA. 

Color,  deep  green  with  secondary  floccose  masses  of  mycelium  in  some  cultures; 
reverse  and  color  in  media,  red  to  very  dark  red,  or,  colorless  in  certain  media. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  thin,  widespread  but  characteristic  growth;  lique- 
faction, none  or  very  slight;  litmus  reaction  acid.    Potato  agar  and  bean  agar,  typical 


PENICILLIUM  DECUMBENS. 


71 


colonies  with  red  color  in  medium.  Potato  plugs,  typical,  reverse  of  colony  and  potato 
both  deep  red.  Raulin's  fluid,  good  growth,  but  no  color  in  fluid.  Cohn's  solution, 
germination  only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar  3  per  cent,  good  growth,  but 
no  red  color.  Lactose  3  per  cent,  very  little  growth.  Lactic  acid  0.9  per  cent,  little 
growth.  Levulose  3  per  cent,  little  growth.  Galactose  3  per  cent,  little  growth. 
Glycerin  3  per  cent,  germinated  only,  grew  when  sugar  was  added.  Potato  starch  3 
per  cent,  good  colonies,  but  no  red  color.  Butterfat,  rather  small  colonies  with  many 
delicate  coremia,  reverse  of  colonies  red,  with  no  color  in  fluid. 

Milk,  good  growth  with  scattered  coremia  in  old  cultures;  curdling  (0.25  per  cent 
calcium  chlorid  added)  very  slow — about  4  weeks;  digestion  slow  or  slight,  no  clear 
fluid;  color  in  milk,  colony  deep.red  below,  milk  deep  red  (vinosus)  at  top,  shading  to 
white  below,  very  slowly  colored. 

Grew  about  equally  well  at  37°  C.  and  20°  C. 

PENICILLIUM  DECUMBENS  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  pura  vel  agaro  Solani  tuberosi  aut  phaseoli 
cultis,  griseo-glaucis,  griseis,  demun  brunneolis,  sparsis;  in  saccharo  oflScinarocommixto 
densior,  glaucescentibus;  parte  aeria  ex 
hyphis  decumbentibus  vel  stoloniformibus 
conidiophoros  brevissimos  gerentibus, 
denum  caespitulis  albis  densis  hypharum 
sterilium  secundariarum,  conspersis ;  re- 
verso  incolorato;  conidiophoris  20-100 X3/x, 
basidiis  7-9X2-3/£,  in  imo  verticillo  denso 
gerentibus;  fructibus  conidicis  ex  catenis 
conidiorum  primum  in  columno  usque  100/z 
longo,  mox,  in  capitulo  conglutinato  solutis; 
conidiis  globosis,  2.5-3//,  vacuolatis,  Isevi- 
bus,  primum  pallide  glaucis  demum  brun- 
neolis; coloniis  gelatinam  non  liquefa  cien- 
tibus,  alkalinis  lacmo,  saccharophilis, 
odorem  in  saccharo  evolventibus. 

Communicavit,  Prof.  P.  H.  Rolfs,  Miami, 
Fla.,  1905. 

Cultivated  in  gelatin  or  potato  agar,  white 
to  gray,  gray-green  ultimately  yellowish 
brown,  green  in  cultures  with  cane  sugar, 
surface  growth  consisting  of  trailing  or  stolon- 
like hyphse  sparsely  developed  and  so  close 
to  the  substratum  as  to  appear  only  as  fer- 
tile hyphse,  bearing  the  conidiophores  as 
short  branches  20-100/^  in  length,  in  old 
colonies  with  dense  tufts  of  sterile  second- 
ary mycelium  scattered  upon  the  surface; 
conidial  fructifications  consisting  of  single 
verticils  of   crowded    conidiiferous    cells, 

7-9  by  2-3/i,  bearing  conidial  chains  first  in  loose  columns  up  to  100/t  in  length  but  soon 
becoming  enveloped  and  broken  up  in  the  drops  of  fluid  secreted  abundantly  from  the 
mycelium  (Gliocladium-like);  conidia  globose  2.5-3/i,  vacuolate,  smooth,  pale  green 
then  brownish  in  mass;  colonies  do  not  liquefy  gelatin;  give  a  weakly  alkaline  reac- 
tion to  litmus;  produce  a  definite  odor  in  cultures  containing  cane  sugar. 

Contributed  by  Prof.  P.  H.  Rolfs  from  Miami,  Fla.,  1905. 


=-«.-.v«u  -'<e£.;«"=>!5;7^--=«<- 


FiQ.  28.— PeniciUium  decwmberu:  a,  b,c,d,  conidial 
fructification,  a  single  verticil  of  conidiiferous 
cells  ( X  900);  ft,  j,  it,  sketches  of  conidial  fructifi- 
cations, with  diagram  of  habit  and  appearance 
of  young  culture  on  potato-agar  (X  140). 


72  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

CULTURAL  DATA. 

Color  gray,  gray-green,  often  gray  or  gray-brown  when  old;  reverse  white;  color  in 
media,  none. 

Odor,  distinct  in  cane-sugar  media. 

Fifteen  per  cent  gelatin  in  water,  medium  growth,  gray -green  to  brown  when  old; 
liquefaction,  none;  litmus  reaction  neutral.  Potato  agar  and  bean  agar,  rather  small 
colonies,  weak  growth,  grayish  green  to  yellow-brown.  Potato  plugs,  white  to  yellow- 
ish brown  colonies,  very  weak  growth.  Raulin's  fluid,  rich  growth,  bright  green,  dis- 
tinct odor.     Cohn's  solution,  germination  only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  rich  growth  up  to  30  per 

cent.     Lactose  3  per  cent,  slight  growth.     Lactic  acid  0.9  per  cent,  medium  colony, 

light  green.     Levulose  3  per  cent,  small  greenish  colonies.    Galactose  3  per  cent, 

growth,  faintly  alkaline  reaction.    Glycerin  3  per  cent,  germination  only.     Potato 

.starch  3  per  cent,  very  slight  growth.     Butterfat,  weak  colonies. 

Milk,  not  adapted  to  this  species,  colonies  grow  very  slowly;  curdling  (0.25  per  cent 
calcium  chlorid  added)  slow — about  4  weeks;  digestion,  little  or  none;  color  in  milk, 
none. 

At  37°  C.  some  growth;  check  at  20°  C.  better  than  37°  C. 

PENICILLIUM  DIVARICATUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  phaseoli  cultis,  avellaneis,  nim- 
quam  viridibus,  in  substrato  late  crescentibus;  parte  aria  ex  hyphis  fertilibua  intri- 
catis,  demum  fere  pulverulenta;  reverse  incolorato;  hyphis  fertilibus  septatis,  ple- 
rumque  brevibus,  repentibus  vel  adscendentibus;  fructibus  conidicis  aut  terminal!  bus 
aut  lateralibus  ex  hyphis  fertilibus  repentibus  ex  verticillis  sessiles  ramorum  et  basid- 
iorum,  irregulariter  in  hyphis  fertilibus  orientibus;  basidiis  15-20X3//,  sterigmatibus 
longis  acuminatis,  in  baside  confertis,  apice  late  divergentibus,  catenas  longas  coni- 
diorum  gerentibus;  conidiis  ellipticis  vel  fusiformibus,  5-7X2. 5-3/t,  avellaneis,  lO/x 
incrassatis  2-3  tubis  germinantibus;  coloniis  gelatinam  non  liquefacientibus,  alka- 
linis  lacmo. 

Legit,  C.  Thom,  Storrs,  Conn. 

Cultivated  in  gelatin  or  bean  agar,  yellowish  brown  (avellaneous),  never  green, 
broadly  spreading  in  the  substratum;  superficial  growth  consisting  only  of  closely 
woven  fertile  hyphse,  becoming  powdery  in  appearance  when  mature;  reverse  of  colony 
not  discolored;  fertile  hyphse  septate,  usually  short,  mostly  creeping;  conidial  fructi- 
fications either  terminal  or  on  short  branches  of  creeping  or  partially  erect  hyphae,  con- 
sisting of  separate  conidiiferous  cells,  of  verticils,  or  of  series  of  verticils  of  branchlets 
and  conidiiferous  cells  irregularly  distributed  along  the  fertile  hyphse;  conidiiferous 
cells  15-20  by  3;^,  with  long  acuminate  sterigmata,  broadly  divergent  at  the  apices 
and  bearing  long  chains  of  conidia;  conidia  elliptical  or  fusiform,  5-7  by  2.5-3/i,  yel- 
lowish to  brownish,  swelling  in  germination  to  lOfi  and  producing  2  or  more  tubes; 
does  not  liquefy  gelatin;  litmus  reaction  alkaline. 

Unmistakable  when  once  seen  in  culture.  Found  in  a  mucilage  bottle,  Storrs, 
Conn.,  1904.     Later  contributed  by  Prof.  G.  F.  Atkinson  from  North  Carolina. 

In  common  with  several  other  forms  included  in  the  genus  Peni- 
ciUium,  the  conidial  fructifications  of  this  species  are  not  strictly  peni- 
cillate  and  terminal.  Every  gradation  is  found  from  fruiting  s^'^stems 
typical  of  the  genus  to  simple  chains  of  conidia  borne  by  single  cells  or 
basidia  upon  prostrate  or  even  submerged  hyphas.  It  partakes,  how- 
ever, of  the  cultural  character  of  the  species  of  the  genus,  as  shown  by 
its  copious  growth  upon  many  different  substrata. 


PENICILLIUM  LILACINUM. 


73 


CULTURAL   DATA. 

Color  light  clay  to  chocolate  or  yellow  (avellaneous,  nearly)  to  darker,  approaching 
brownish  yellow,  never  green ;  reverse  uncolored;  color  in  media,  none. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  good  growth;  liquefaction,  none;  litmus  reaction, 
acid.  Potato  agar,  bean  agar,  and  potato  plugs,  typical.  Raulin's  fluid,  typical. 
Cohn's  solution,  germination  only. 

Synthetic  fluid  (Dox's)  carbon  supplied  as:  Cane  sugar,  good  growth  up  to  30  per 
cent,  acid  reaction.  Lactose  3  per  cent,  slight  growth.  Lactic  acid  0.9  per  cent, 
medium  growth,  not  fully  normal.     Levulose  3  per  cent,  small  development.    Galac- 


FiQ.  29.—Penicillium  divaTtcatum:  a,  d,  e,  f,  conldiiferous  cells,  conidia  and  their  arrangement  (X  1,600); 
6,  c,  irregular  types  ol  arrangement;  ^,  A,)!:,  sketches  of  conidial  fructification  (X  200);  m,  n,  germination 
of  conidia  (X  900). 

tose  3  per  cent,  slow  growth,  reaction  neutral.  Glycerin  3  per  cent,  germinated  only; 
grew  when  sugar  was  added.  Potato  starch  3  per  cent,  typical.  Butter  fat,  weak 
growth,  but  characteristic  fruiting. 

Milk,  slow  and  weak  colonies;  curdling  (0.25  per  cent  calciiun  chlorid  added)  slow — 
14  days  or  more;  digestion,  slow  and  slight;  color  in  milk,  none. 

Colonies  grew  better  at  37°  C.  than  at  20°  C. 

PENICILLIUM  LILACINUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  phaseoli  cultis,  albis,  vel  albis  demum 
pallide  lilacinis  imprimis  in  saccharo  officinaro  commixto,  floccosis;  hyphis  aereis 
ramosis,  adscendentibus,  septatis,  Zfi  cr.,  ramos  fertiles  brevissimis  gerentibus;  reveiso 


74 


CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 


incolorato;  fructibus  conidicis  usque  100  longis,  e  basidiis  sessilibue,  eolitariis  vel  verti- 
cillatis,  aut,  e  ramia  brevissimia  vel  apicibua  hyphorum  aeriorum,  1,  2,  3  verticillos 
ramulorum  et  basidiorum,  catenas  longas  et  divergentea  conidiorum  gerentum;  baaidiis 
basidibus  incrassatis,   apicibua  acuminatis  et  divergentibus,  7-10  longis;  conidiis 
2.5-3X2/1  ellipticia,  laevibus,  pallide  lilacinis. 
Coloniis  gelatinam  lente  liquefacientibus,  alkalinis  lacmo. 
Comm.,  Prof.  G.  F.  Atkinaon  et  C.  W.  Edgarton,  Ithaca,  N.  Y. 
Cultivated  in  pure  gelatin  or  bean  agar  white,  white  to  pale  lilac  in  cultures  con- 
taining augara,  more  or  less  loosely  floccose  with  hyphae  branched,  aeptate,  ascending,  3fi 
in  diameter,  producing  conidial  masses  upon  very  short  branchea  irregularly  diatrib- 
uted,  or  becoming  conidiophorea  toward  the  apex;  reverae  of  colony  not  discolored; 
conidial  fructificationa  up  to  100/i  in  length,  consisting  of  solitary,  aeaaile  conidiiferous 
cella,  or  verticils  of  conidiiferous  cells,  or  short  branches  bearing  1,  2,  or  3  verticils  of 
branchleta  and  conidiiferous  cells  with  long,  tangled  chains  of  conidia.     Conidiiferous 

cells  flask-shaped,  divergent  at 
the  apices,  acuminate,  7-10/t  in 
length ;  conidia  elliptical ,  amooth, 
2.  5-3  by  2/i,  thin  walled,  pale 
lilac.  Colonies  slowly  liquefy 
gelatin,  with  strongly  alkaline 
reaction. 

Received  from  Prof.  G.  F.  At- 
kinson and  C.  W.  Edgarton, 
Ithaca,  N.  Y. 


A  relationship  of  this  spe- 
cies to  the  common  green 
forms  is  very  doubtful.  The 
chains  of  conidia  produced 
break  up  so  quickly  and 
completely  in  moimting  in 
fluid  for  examination  that  it 
is  often  difficult  to  find  even 
a  single  conidium  attached 
to  its  sterigma.  The  hyphse 
with  branches  and  basidial 
cells,  aside  from  the  produc- 
tion of  long  conidial  chains, 
might  readily  be  placed  in  any  one  of  several  hyphomycete  genera. 
The  form  of  conidial  fructification  varies  from  a  single  conidiiferous 
cell  or  basidium  with  a  chain  of  conidia  upon  an  aerial  hypha  to  a 
single  verticil,  or  a  branch  with  two  or  three  successive  verticils  and 
even  to  a  terminal  fructification  allying  it  with  the  typical  penicillate 
forms. 

CULTURAL   DATA. 

Color  white  to  a  characteristic  lilac  shade;  reverse  of  colony  white;  color  in  media, 
none. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  fair  growth,  not  heavy,  white;  liquefaction 
rather  slow — 14-16  days;  litmus  reaction  strongly  blue     Peptone  milk  sugar  gela- 


FiQ.  SO.—PeniciUium  lilacinum:  a,  h,  c,  short  conidiophores 
and  verticils  of  conidiiferous  cells  showing  the  various  branch- 
ing and  arrangement  of  cells  (X  1,000);  d,  conidiiferous  cell, 
solitary  and  sessile  on  an  aerial  hypha,  not  uncommon  in 
this  species  (X  1,600);  e,  conidia  (X  1,000);  /,j?,  ft, sketches  of 
conidial  fructifications,  varjnng  from  a  single  chain  to  a  typi- 
cal penicillate  form  (x  260). 


PENICILLIUM   INTRICATUM. 


75 


tin  (Conn's),  liquefied  in  2  weeks,  white  colonies.  Potato  plugs,  white  colony; 
Cohn's  solution,  very  weak  growth  but  typical  lilac  color. 

Synthetic  fluid  (Dox's),  carbon  applied  as:  Cane  sugar  3  percent  to  30  p>er  cent, 
good  growth  with  typical  lilac  color,  alkaline  reaction,  no  fermentation.  Lactose  3 
per  cent,  germination  only,  slight  growth.  Lactic  acid  0.9  per  cent,  germination  only. 
Levulose  3  per  cent,  slow  growth.  Galactose  3  per  cent,  slow  development  with 
alkaline  reaction.  Glycerin  3  per  cent,  not  characteristic,  little  more  than  ger- 
mination. Butterfat,  typical  colony  giving  brownish  color  to  fluid  and  causing  drops 
of  yellow  oil  to  separate  out. 

Milk,  curdling,  slow;  digestion,  slow;  color  in  milk,  none. 

At  37°  C,  best  growth;  at  20°  C,  good  growth. 

PENICILLIUM  INTRICATUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatina  vel  agaro  phaseoli  cultis,  albis,  griseis,  griseo- 
glaucis,  demum  griseis,  lente  fere  fuligineis,  floccosis;  zonatis;  parte  aeria  usque  1-3 
mm.  cr.,  exhyphis  aereis  ramosis 
dense  intricatis;  reverso  incolo- 
rato  vel  sulphureo  interdum 
lente  avellaneo;  substrato  sul- 
phureo colorato;  conidiophoris 
interdum  terminalibus  plerum- 
que  ex  hyphis  a«reis  brevibus 
30-50/i  ramosis;  fructibus  conid- 
icis  50-100/t  usque  140fi  longis — 
multo  longioribus  in  substratis 
saccharinis — ex  verticillo  basidi- 
orum,  vel  ex  1-3  verticillis  ba- 
sidiorum  in  ramis  divergenti- 
bus,  vel  ex  verticillis  ramulorum 
et  basidiorum  eodem  verticillo, 
catenis  conidiorum  saepe  co- 
lumno  laxe  convergentibus;  ba- 
sidiis  8-10X2-2.5//,  pauci8(4-10) 
verticillo,  cum  catenis  basidi- 
orum divergentibus;  conidiis  el- 

lipticisvelglobosis,  hyalinis,  velpallideglaucis,  2.5-3/tdiam.,  laevibus,  leptodermibus, 
intus  granulosis,  in  catenis  manentibus  submersis;  coloniis  gelatinam  nonliquefacien- 
tibus,  alkalinis  lacmo. 

Culturae  ex  humo,  Prof.  W.  M.  Esten,  Storrs,  Conn.,  1907. 

Colonies  upon  gelatin  or  bean  agar,  white,  gray,  greenish  gray,  when  old  gray  or 
smoky,  floccose,  becoming  a  mass  of  interwoven  hyphse  and  ropes  of  hyphae  1-3  mm. 
in  thickness;  reverse  of  colony  and  substratum  not  colored  in  bean  agar,  more  or  less 
sulphur  yellow  or  even  bro^vnish  in  sugar  media;  conidiophores  sometimes  terminal, 
more  commonly  branches  of  aerial  hyphse  30-50/Lt  in  length;  conidial  fructifications 
50-100  up  to  140/1  in  length,  or  much  longer  in  old  sugar  cultures,  consisting  of  simple 
verticils  of  conidiiferous  cells,  or  of  1-3  verticils  upon  divergent  branchlets,  or  of 
branchlets  and  conidiiferous  cells  in  the  same  verticil;  conidiiferous  cells  8-10  by 
2-2. 5/i,  few  (4  to  10)  in  each  verticil,  bearing  more  or  less  divergent  chains  of  conidia 
frequently  aggregated  into  a  loose  column;  conidia  elliptical  or  globose,  hyaline  or 
pale  greenish,  2.5-3/t  diameter,  smooth,  thin  walled,  granular  within,  remaining  in 
chains  in  fluid  mounts^  colonies  alkaline  to  litmus,  not  liquefying  gelatin. 

Found  in  cultvu-es  from  soil,  Storrs,  Conn.,  by  Prof.  W.  M.  Esten,  1907. 


Fig.  31. — Pfnicillium  ivtricatuin :  a,  b,  c,  conidial  fructification, 
conidiiferous  cell,  conidial  chain  (x  l,(tX));  d,  c,/,  sketches  of 
conidiophores,  branching  and  arrangement  (X  200). 


76 


CULTURAL  STUDIES  OP  SPECIES  OP  PENICILLIUM. 


CULTURAL  DATA. 

Color  white  or  greenish  gray — not  green,  grayish  to  brown  or  drab;  reverse  and 
medium  uncolored  or  sulphur-yellow  in  some  media;  litmus  reaction  strongly  alka- 
line. Potato  and  bean  agar,  typical.  Potato  plugs,  weak  growth,  not  adapted  to 
this  species.     Cohn's  solution,  weak  growth,  yellowish-green  colonies. 

Synthetic  fluid  (Dox's)  carbon  supplied  as:  Cane  sugar,  grew  well  up  to  30  per  cent. 
Lactose  10  per  cent,  good  colonies.  Levulose  3  per  cent,  good  growth,  yellowish 
mycelium,  fluid  yellowish,  alkaline.  Galactose  3  per  cent,  typical  alkaline  reaction. 
Glycerin  3  per  cent,  slow  growth.  Butterfat,  good  growth,  reverse  yellow,  fat  little 
changed. 

Milk,  fruiting  areas  upon  glass,  mycelium  in  contact  with  milk  sulphur-yellow; 
curdling  (0.25  per  cent  calcium  chlorid  added)  very  slow — 3  weeks;  digestion  very 
slow  in  normal  or  acid  milk,  rapid  when  alkaline;  color  in  milk,  none. 

At  37°  C.  grew  more  rapidly  than  at  20°  C. 


PENICILLIUM  SPINULOSUM  n.  sp. 

Latin  diagnosis. — Coloniis  in  gelatinavel  agaro  phaseoli  cultis, atro-viridibus,  demum 
fere  atris,  cito  et  late  in  substrate  crescentibus,  margine  sterili  lata  juvenilibus;  parte 

aeria  ex  conidiophoris 
et  ex  hyphis  floccosis 
sparsis  composita ;  re- 
verso  incolorato ;  conidio- 
phoris 105-300X3-3.5;/, 
vel  longioribus,  apice 
5fi  incrassato,  verticillum 
basidiorum  9.5-11X2-3/Z 
gerente;  fructibus  coni- 
dicis  in  columno  denso 
300  usque  500X15-30/1 
ex  catenis  conidiorum 
compositis ;  conidiis  pyri- 
formibus  vel  globosis, 
3.2-3.5X3.6-4;(,  lepto- 
dermibus,  primum  laevi- 
bus  demum  ministissime 
spinulosis;  coloniis  gela- 
tinam  lente  liquefacien- 
tibus,  acidis  lacmo. 

In  cultura  in  labora- 
torium,  Hannover,  Ger- 
mania. 

Cultivated  upon  gelatin  or  bean  agar,  deep  green,  spreading  broadly  in  the  sub- 
stratum with  broad  sterile  margin  when  young;  aerial  portion  consisting  of  conidio- 
phores  and  scattered  aerial  hyphaj;  reverse  of  colony  not  discolored;  conidiophorea 
150-300;u  or  longer  by  3-3. 5ju,  with  apex  enlarged  to  5/i  in  diameter,  bearing  a  single 
verticil  of  conidiiferous  cells  9.5-11  by,2-3/i;  conidial  fructification  a  close  column  of 
conidial  chains  up  to  300  or  even  500/t  in  length  by  15-30,u;  conidia  pyriform  to  glo- 
bose, 3.2-3.5  by  3.6— 4/i,  very  thin  walled,  smooth  at  first  then  delicately  spinulose 
or  verrucose,  yellowish  green  then  almost  smoky;  liquefying  gelatin  slowly,  with 
strongly  acid  reaction. 

Found  as  a  contamination  of  another  species  of  Pcmcillium  obtained  in  Doctor 
Wehmer's  laboratory  at  Hanover,  Germany.     Easily  recognized  and  cultivated. 


Fio.  .'i2. — Penicillium  spinulosum:  a,  ft,  conidial  fructifications  consisting 
of  single  verticils  of  conidiiferous  cells  (X  900);  c,  conidiiferous  cell  with 
chain  of  young  conidia  smooth  (X  900);  d,f,  ripe  conidia,  delicately 
echinulate  (X  900);  e,  swollen  end  of  conidiophore  bearing  conidiiferous 
cells  (X  900);  g,  h,  sketches  of  conidial  fructifications  (X  1,400). 


PENICILLIUM   NO.   28. 


77 


CULTURAL   DATA. 

Color  deep  dull  green;  reverse  cream,  or  slight  traces  of  pink  or  violet;  color  in 
media,  none. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  slow  but  typical;  liquefaction,  rather  slow  and 
variable;  litmus  reaction,  acid.  Potato  and  bean  agar  and  potato  plugs,  typical, 
producing  a  very  heavy  layer  of  dark-green  conidia  when  cane  sugar  is  added.  Rau- 
lin's  fluid,  typical.    Cohn's  solution,  germination  only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  grows  well  up  to  50  per 
cent  solution  with  acid  reaction.  Lactose  3  per  cent,  weak  growth.  Levulose  3  per 
cent,  typical.    Glycerin  3  per  cent, 

f 


Potato  starch 
Butterfat,  rich 


half  normal  growth 
3  per  cent,  typical, 
growth. 

Milk,  colonies  grow  very  slowly; 
curdling  (0.25  per  cent  calcium 
chlorid  added)  very  slow;  digestion, 
very  slight;  color,  none. 

At  37°  C.  grew  better  than  check 
at  20°  C. 

PENICILLIUM  No.  28. 


Colonies  upon  sugar  gelatin  and 
potato  or  bean  agar,  gray-green  with 
broad  white  border  when  growing, 
floccose,  tangled  tufts  of  hyphse 
and  ropes  of  hyphae  spreading  inde- 
terminately upon  the  substratum, 
reverse  yellow  or  tan  on  media  con- 
taining sugar,  conidiophores  arising 
direct  from  the  substratum  as  short 
lateral  branches  from  38-160/i  in 
length,  3/i  in  diameter,  swelling  to 

5/i  at  apex,  from  aerial  hyphae  or  ropes  of  hyphae.  Conidial  fructification  a  simple 
column,  300/i  or  even  500/i  in  length  by  10-15//  in  diameter,  produced  from  a  single 
whorl  of  conidiiferous  cells  at  the  apex  of  the  conidiophore.  Conidia  elliptical  to 
globose,  2-3/ior  2-2.4//  by  3-3. 3/i  in  diameter,  light  yellowish  green  in  mass,  smooth. 
Colonies  liquefy  sugar  gelatin.    Litmus  reaction  strongly  acid. 

Grows  readily  upon  all  common  media. 

Found  at  Storrs,  Conn.,  upon  decaying  mushroom.     Very  characteristic  and  read- 
ily recognized   from  others  with  related  morphology. 


Fig.  33.—PenicilUum  No.  28:  o,  6,  conidial  fructifications 
each  a  single  verticil  (X  900);  g,  h,  j,  sketches  of  conidio- 
phores and  fructification  (X  140);  fc,  tip  of  conidiophore, 
swollen  at  apex,  bases  of  lowest  two  conidiiferous  cells 
(X  1,600). 


CULTURAL  DATA. 

Color  gray-green;  reverse  yellow,  or  tan  when  sugar  is  present;  color  in  media, 
more  or  less  yellow,  according  to  media. 

Fifteen  per  cent  gelatin  in  water,  good  growth,  clear  green;  liquefaction  slow — 2 
weeks  or  more;  litmus  reaction  acid.  Potato  agar,  typical  growth,  acid  reactions. 
Bean  agar,  typical  growth,  acid  reactions.  Raulin's  fluid,  good  growth,  edges  pink, 
fluid  brownish  fluorescent.     Cohn's  solution,  weak  growth. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  good  growth  up  to  50 
per  cent,  acid  reaction.     Lactose  3  per  cent,  slow  but  fairly  typical  growth.    Lactic 


78  CULTURAL  STUDIES  OF  SPECIES  OF  PENICILLIUM. 

acid  0.9  per  cent,  fair  growth,  not  good.  Levulose  3  per  cent,  good  but  slow  colo- 
nies, acid  reactions.  Glycerin  3  per  cent,  medium  growth,  pink  tinge  to  fluid.  Potato 
starch  3  per  cent,  good  growth,  yellow  drops  of  transpired  fluid,  fluid  tinged  yellow 
at  top  only.     Butterfat,  rich  growth,  fluid  reddish  brown. 

Milk,  good  growth,  acid  in  litmus  milk;  curdling  (0.25  per  cent  calcium  chlorid 
added)  in  8-9  days,  good  curd;  digestion  slow,  incomplete;  color,  pale  yellow  in 
digested  fluid. 

At  37°  C,  killed;  check  at  20**  C,  good. 

SPECIES  FORMING  PINK  SCLEROTIA. 

Four  races  have  been  found  in  which  pink  sclerotia  are  regularly 
formed  in  culture.  These  sclerotia  are  elliptical  to  globose  and 
from  200  to  500/^  in  diameter.  They  begin  to  be  formed  within  the 
first  week  in  richly  nourished  cultures.  Although  examined  repeat- 
edly, no  trace  of  ascus  formation  has  yet  been  found.  These  forms 
are  included  here  under  their  serial  numbers,  29,  30,  31,  and  32, 
rather  than  with  specific  names.  The  descriptions  and  figures 
introduced  will,  it  is  thought,  identify  these  organisms  clearly  in 
their  penicilhum  form,  but  the  uniformity  of  sclerotium  production 
makes  ascus  production  so  probable  under  proper  conditions  that 
it  seems  best  not  to  give  specific  names  to  this  imperfect  form  when 
some  of  them  may  be  already  recognizable  by  others  or  by  further 
investigation. 

PENicnxiuM  No.  29. 

Colonies  grown  upon  gelatin  and  potato  or  bean  agar  white  to  gray-green,  sometimes 
partly  clear  green,  becoming  zonate  with  rings  of  pink  sclerotia  in  age,  sparsely  or 
loosely  floccose,  indeterminate  broadly  spreading  margins  persistently  white,  slightly 
yellow  below.  Conidiophores  80-200/x  or  even  400/i  by  3-5/x  commonly  150-200/i 
in  length  as  branches,  usually  perpendicular,  from  hyphse  4-5^  in  diameter.  Co- 
nidial  fructification  a  single  verticil  of  rather  few  (about  12-15)  conidiiferous  cells 
9  by  2fi,  producing  chains  of  conidia  in  a  loose  column  150-250/i  or  even  400/z  by  20- 
30jtt.  Conidia  elliptical,  3-3.6  by  2.3-2.8/i,  smooth,  very  pale  blue  (transmitted 
light).  Sclerotia  in  loose  networks  of  mycelium,  numerous,  pink,  elliptical  to  globose, 
150-300/£  in  diameter.  These  begin  to  appear  in  one  week  in  gelatin  cultures.  Colo- 
nies liquefy  sugar  gelatin  slowly  but  completely  in  10-12  days.  Give  a  strong  acid 
reaction  with  litmus  media. 

Characterized  by  the  production  of  large  numbers  of  pink  sclerotia  with  compar- 
atively small  quantities  of  conidia,  whereas  the  next  form  (Penicillium  No.  30)  pro- 
duces few  sclerotia  and  great  quantities  of  conidia. 

Collected  at  Storrs,  Conn.,  on  decaying  mushroom.  Probably  not  closely  related 
to  the  common  species  of  Penicillium,  but  its  occurrence  in  culture  and  ready  adap- 
tation to  all  media  tried,  in  numerous  cultures,  justify  its  inclusion  with  these  species. 

CULTURAL  DATA. 

Color  white  to  gray  or  green,  with  many  pink  sclerotia;  reverse  colorless  or  slightly 
salmon;  color  in  media  slightly  yellow  in  some  media. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  good  growth;  liquefaction,  15  days;  litmus  reac- 
tion alkaline.  Potato  agar  and  bean  agar,  typical  colonies,  white  or  gray,  with  few 
green  areas  and  abundant  pink  sclerotia.     Raulin's  fluid,  typical.    Colonies  upon 


SPECIES  FORMING  PINK  SCLEBOTIA. 


79 


media  with  cane  sugar  produce  sclerotia  more  numerously  and  more  quickly  (5  days) 
than  without  sugar.     Butterfat  as  a  source  of  carbon  in  Dox's  fluid,  typical  colonies. 
Milk,  typical;  curdling  (0.25  per  cent  calcium  chlorid  added)  in  9  days;  digestion, 
Blow;  color  in  milk,  none. 

Penicillium  No.  30. 

Colonies  on  sugar  gelatin  and  potato  or  bean  agar  gray-green  or  green  persistently, 
Bin-face  growth  mostly  of  crowded  conidiophores  and  producing  pink  sclerotia  at  the 
Burface  or  partly  embedded  in  the  substratum  150-300/t  in  diameter,  broadly  spreading. 
Conidiophores  from  240-525/(,  usually  about  SOO/i,  in  length  either  arising  separately 
from  the  substratum  or  as  branches  very  close  to  its  surface.  Conidial  fructification 
a  single  verticil  of  conidiiferous  cells  bearing  conidia  in  a  close  column  up  to  500/«  in 
length  by  15-30/t.  Conidia  elliptical  or  subglobose  2.5  by  3/i  or3/t,  withaslight  green- 
ish color.  Colonies  liquefy  sugar  gelatin  rather  slowly,  and  give  an  acid  reaction 
with  litmus. 


Fig.  34. — Penicillium  No.  29:  a,  b,  conldiopliore  and  verticil  of  conidiiferous  cells  (X  900);  c,  d,  e,  germina- 
tion of  conidia  (X  900);  /,  ;;,  A,  j,  sketches  of  conidial  fructifications  (X  140);  fc,  diagrammatic  sketch 
from  photomicrograph  showing  relations  of  sclerotia  and  conidial  fructifications. 

Apparently  related  to  No.  29,  but  differing  in  the  length  and  density  of  its  column 
of  conidia,  in  the  position  of  the  sclerotia,  in  habit,  in  culture,  and  in  its  acid  reaction. 
Collected  at  Storrs,  Conn.,  upon  decaying  LaciaritLS  vellereas,  September,  1904. 


CULTURAL   DATA. 

Color  green  or  grayish  green,  persistently,  with  abundant  pink  sclerotia;  reverse 
uncolored;  color  in  media,  none. 

Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  typical;  liquefaction  rather  slow;  litmus  reac- 
tion acid.  Potato  agar  and  bean  agar,  typical.  Potato  plugs,  typical.  Cohn's 
solution,  germination  only. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  rich  growth.  Lactose  3 
per  cent,  slowly  typical  colonies,  acid  reaction.  Lactic  acid  0.9  per  cent,  weak 
growth.  Levulose  3  per  cent,  medium  growth.  Galactose  3  per  cent,  typical.  Gly- 
cerin 3  per  cent,  very  weak  growth.    Butterfat,  slow,  but  characteristic  colonies. 

Milk,  curdling  (0.25  per  cent  calcjum  chlorid  added)  in  9  days;  digestion,  slow  but 
complete;  color  in  milk,  none. 

At  37°  C.,  no  growth;  grew  when  cooled;  check  at  20°  C,  typical. 


80 


CULTURAL  STUDIES   OF   SPECIES  OF   PENICILLIUM. 


Peniciluum  No.  31. 

Colonies  upon  gelatin  and  potato  or  bean  agar  from  white  to  gray  to  gray-green 
mostly  white,  with  few  areas  of  green  conidia  sprinkled  with  pink  sclerotia,  sparsely 
floccose,  broadly  spreading.  Conidiophores  branching  from  aerial  hyphae,  very  short 
to  380/i  in  length,  commonly  150-240/1,  conidial  fructification  with  a  single  verticil 
or  once  branched  with  branch,  conidiiferous  cells  and  chains  of  conidia  divergent,  up 
to  140/i  in  length,  but  usually  much  less.  Conidia  2.5-3;x  globose,  smooth,  rarely 
found  in  quantity  to  color  the  colony.  Sclerotia  elliptical  or  globose,  160-330//,  pink, 
developed  in  10-15  days.  No  asci  have  been  secured.  Colonies  liquefy  sugar  gelatin 
rapidly  and  give  a  stongly  alkaline  reaction  to  litmus  in  the  same  cultures. 
Grows  readily  in  conidial  transfers  upon  all  common  media. 

Collected  upon  decaying  Clavaria  at  Storrs,  Conn.,  September,  1904.     Identical 

culture  sent  from  Cambridge,  Mass.,  by 
Dr.  A.  F.  Blakeslee  in  culture  obtained 
from  fruit  imported  from  Porto  Rico. 

CULTURAL  DATA. 

Color  white  or  gray,  conidial  areas  gray- 
green,  very  numerous  pink  sclerotia;  re- 
verse colorless  or  with  yellow  areas;  color 
in  media,  none  or  slightly  yellowish. 
Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  typ- 
ical white  or  gray  colonies;  liquefaction 
rapid;  litmus  reaction  alkaline.  Potato 
and  bean  agar,  typical,  cultures  with  sugar 
added  become  distinctly  greener  than 
others.  Potato  plugs,  typical,  white  or 
gray  with  greenish  areas,  sclerotia,  and 
crystal  drops  of  transpired  fluid.  Rau- 
lin's  fluid,  some  growth,  not  entirely 
typical.  Cohfl's  solution,  weak  develop- 
ment but  characteristic. 

Synthetic  fluid  (Dox's),  carbon  sup- 
plied as:  Cane  sugar  1.5-20  per  cent,  typ- 
ical growth.  Lactose  3  per  cent,  weak  growth.  Lactic  acid  0.9  per  cent,  no  growth. 
Levulose  3  per  cent,  typical.  Galactose  3  per  cent,  typical,  alkaline.  Glycerin  3 
per  cent,  slight  growth.     Butterfat,  typical  growth. 

Milk,  ciu-dling  (0.25  per  cent  calcium  chlorid  added)  in  9  days;  digestion  com- 
plete; color,  none. 
At  37°  C,  no  growth,  grew  when  cooled;  check  at  20°  C,  typical. 

Peniciluum  No.  32. 

Colonies  upon  milk-sugar-gelatin  and  potato  or  bean  agar  gray-green;  floccoee,  but 
with  aerial  part  mostly  long  conidiophores  and  few  vegetative  hyphae,  slightly  yel- 
lowish to  pronounced  salmon  color  below;  broadly  spreading;  developing  elliptical  to 
globose  sclerotia  150-200/t  in  diameter  at  the  surface  of  the  substratum  in  2-3  weeks. 
Conidiophores  200-500/x  by  3-4//.  Conidial  fructification  a  verticil  of  3-5  branches 
10-17/1  by  2-3/z  rarely  a  secondary  verticil,  each  bearing  a  dense  verticil  of  conidiiferous 
cells,  8-10/t  by  2/t  producing  long,  parallel,  or  slightly  divergent  chains  of  conidia. 
Conidia  elliptical  or  fusiform,  3.5-4/t  by  2-3//,  green,  granular  within,  smooth,  swelling 
in  germination  to  6//  and  producing  from  one  to  several  germ  tubes.  Colonies  slowly 
liquefy  milk-sugar-gelatin  and  produce  purple  or  neutral  colors  in  litmus  media. 

Sent  by  Prof.  P.  H.  Rolfs  from  Miami,  Fla.,  upon  portion  of  pineapple,  March,  1905. 


FlQ.  35.—Penicillium  No.  31 :  o,  6,  branching  of  conld- 
iophore(X  900);  c,  germination  of  conldia(X  900); 
d,  e,  f,  sketches  of  conidiophores. 


COMPARATIVE   CULTURAL  DATA. 


81 


CULTURAL   DATA. 

Color  gray-green,  with  scattered  white  to  pink  sclerotia;  reverse,  sulphur-yellowish 
to  pronounced  salmon;  color  in  media  reddish  or  yellowish  in  special  cases,  others 
none.    Odor,  none. 

Fifteen  per  cent  gelatin  in  water,  typical;  liquefaction,  none  in  15  days,  later  very 
slow  liquefaction;  litmus  reaction  neutral,  leaves  both  acid  and  alkaline  media 
purple-blue.  Potato  agar  and  bean  agar,  typical,  slightly  thirmer  than  gelatin  cul- 
tures, gray-green  without  sugar,  clear  green  with  cane  sugar.  Potato  plugs,  typical, 
transpires  yellow  drops  which  become  very  dark  yellow  (balsam).  Raulin's  fluid, 
good  colonies  becoming  rosy  below.  Cohn's  solution,  small  colonies,  fluid  slightly 
yellow. 

Synthetic  fluid  (Dox's),  carbon  supplied  as:  Cane  sugar,  grew  in  solutions  up  to  30 
per  cent  with  acid  reaction.  Lactose  3  per  cent,  very  slow  growth  of  small  character- 
istic colonies.     Lactic  acid  0.9  per  cent,  good  growth,  light  green.    Levuloee  3  per 


Fig.  SG.—Penicillium  No.  32 :  o,  6,  d,  branching  of  conidial  fructifications  (a  X  1,400,  b  and  d,  X  900) ; 
c,  a  single  secondary  verticil  (X  1,400);  g,  h,j,  k,  sketches  of  fructifications  of  various  ages 
(X  140);  m,  n,  o,  germination  of  conidia  (X  900). 

cent,  very  slow-growing  but  heavy  colonies.  Galactose  3  per  cent,  typical.  Glycerin, 
very  small  colonies.     Butterfat,  typical  colonies. 

Milk,  curdling  (0.25  per  cent  calcium  chlorid  added)  very  slow;  digestion,  very 
slow;  color  in  milk,  none. 

At  37°  C,  grew  more  rapidly  than  check  at  20°  C. 

COMPARATIVE  CULTURAL  DATA.  ^ 

A  summary  of  accessory  cultural  data  has  already  been  given  for 
each  species  in  connection  with  the  descriptions.  Many  series  of 
cultures  have  been  made  with  numerous  media  to  obtain  data  as  to 
the  ability  of  the  species  studied  to  grow  upon  particular  media  or 
under  particular  conditions.  It  has  been  possible  thus  to  determine 
the  relative  activity  of  single  species  and  groups  of  species. 
Although  particular  species  in  these  cultures  have  shown  unique 
differences  which  assist  in  their  differentiation,  the  most  valuable 
8108— Bull.  118—10 6 


82  CULTURAL  STUDIES   OF   SPECIES  OF   PENICILLIUM. 

result  of  comparative  culture  is  found  in  the  separation  of  the  series 
into  groups  of  races  or  species  which  resemble  each  other  closely  in 
their  metabolic  activities.  Part  of  these  experiments  are  tabulated 
in  Tables  2,  3,  4,  5,  and  6,  and  will  be  discussed  in  the  following 
sections. 

Since  the  complex  composition  of  the  media  in  common  use  for 
cultural  work  makes  the  analysis  of  the  data  obtained  impossible, 
it  was  first  necessary  to  determine  the  reactions  of  these  species  to 
some  of  the  individual  substances  of  which  these  media  are  composed. 
It  has  already  been  noted  that  gelatin  alone  in  distilled  water  sus- 
tains growth  in  the  large  majority  of  the  species  of  this  genus. 
These  cultures  in  certain  species  lack  green  color,  which,  how- 
ever, becomes  present  on  using  the  peptones  and  sugars  added  to 
gelatin  in  most  formulae.  Such  media  are  still  too  complex  to  make 
close  analysis  of  results  possible." 

Many  of  the  determinations  given  were  made  in  duplicate  and  in 
some  cases  the  entire  series  was  repeated  one  to  several  times.  In 
inoculating  cultures  for  this  work  conidia  were  transferred  to  the 
tubes  in  large  numbers,  so  that  their  presence  could  be  detected  by 
examination  with  a  lens.  Where  species  failed  to  grow,  the  doubts 
of  inoculation  were  commonly  dispelled  by  the  addition  of  cane 
sugar,  which  permitted  the  conidia  to  develop  normally  if  present 
and  still  viable.  The  presence  of  germinated  conidia  upon  the  sur- 
face of  a  medium  is  good  evidence  of  proper  inoculation.  The  data 
given  are  believed,  therefore,  to  represent  with  a  fair  degree  of 
accuracy  the  comparative  cultural  reactions  of  the  species  used. 

In  reporting  these  series  of  comparative  cultures,  the  data  have 
been  tabulated  as  far  as  possible  for  convenient  comparison  of  the 
relative  activity  of  the  different  forms.  (See  tables  beginning  on 
p.  98.)  The  names  as  far  as  determined  are  given,  together  with  the 
cultural  number,  in  Table  1 .  In  the  remaining  tables  the  numbers 
are  repeated  without  the  names.  In  studying  the  tables  a  reference 
to  cultural  numbers  will  quickly  locate  the  forms  discussed. 

CULTURES    IN   DISTILLED    WATER. 

To  determine  the  possibilities  of  growth  from  food  stored  m  the 
conidia,  cultures  were  made  in  distilled  water.  Of  forty-four  strains 
imder  cultivation  but  six  showed  clearly  descernible  germination. 
None  produced  more  than  germ  tubes  hanging  down  into  the  fluid. 

a  This  work  was  carried  on  in  cooperation  with  Mr.  A.  W.  Dox,  who  has  studied  the 
metabolism  of  the  species  concerned  with  cheese  ripening  (P.  camemberti  and  P. 
roqueforti),  as  well  as  a  few  other  species,  under  many  conditions  of  culture,  while  the 
writer  has  conducted  comparative  studies  of  a  large  number  of  forms  under  more 
limited  cultural  conditions.  All  chemical  questions  arising  throughout  this  work 
have  been  passed  upon  by  Mr.  Dox. 


COMPARATIVE   CULTURAL  DATA.  83 

AGAR-AGAR    AS    A    SOURCE    OF    FOOD, 

Tubes  of  1 .5  per  cent  agar-agar  in  distilled  water  were  inoculated 
with  eighteen  species  of  Penicillium.  Sixteen  of  these  produced 
growth.  In  all  cases  the  colonies  produced  were  very  small,  some 
of  them  barely  discernible  to  the  naked  eye.  Not  one  of  them  was 
distinctly  colored  by  conidial  masses,  but  in  nearly  every  case  some 
conidial  fructifications  were  found.  These  cultures  show  that  the 
species  tested  were  able  to  obtain  from  the  medium  sufficient  nourish- 
ment for  very  slight  growth. 

AGAR-AGAR  AS  A  SOURCE  OF  CARBON. 

Agar-agar  is  a  carbohydrate  and  might  serve  as  a  source  of  carbon 
if  other  nutrients  were  supphed.  One  and  one-half  per  cent  of  agar 
was  therefore  introduced  into  Mr.  Dox's  synthetic  fluid,  already  con- 
taining all  essential  elements  except  carbon.  Thirty-seven  races  of 
PeniciUium  were  inoculated  into  this  medium,  and  nearly  every 
species  produced  some  growth.  Examined  with  the  microscope, 
conidial  fructifications  were  found  in  nearly  every  case,  but  in  no 
case  was  the  colony  large  enough  or  definite  enough  to  affect  the 
observer's  estimate  of  results  if  such  growth  were  added  to  or  sub- 
tracted from  the  colonies  upon  nutrients  really  adapted  to  sustain 
the  species  studied.  Dox's  stock  solution,  with  or  without  the  addi- 
tion of  agar,  was  in  this  way  shown  to  be  a  safe  medium  for  the  study 
of  the  metabolic  reactions  of  these  species  to  changed  sources  of 
carbon. 

The  possibiHty  of  error  in  the  introduction  of  agar  was  shown  in 
the  following  manner:  One  and  one-half  per  cent  of  agar  was  intro- 
duced into  Dox's  fluid  and  1.1  normal  lactic  acid  added  in  quantity 
to  make  the  whole  0.5  per  cent  acid.  The  medium  was  then  auto- 
claved.  After  this  treatment  the  agar  refused  to  solidify.  Tubes  of 
this  fluid  were  inoculated  with  nine  different  species  of  PeniciUium. 
All  except  P.  hrevicaule  grew  well  and  produced  colonies  recognizable 
by  their  cultural  characters.  In  introducing  agar  in  such  work  it  is 
therefore  necessary  to  guard  against  the  introduction  of  acid  before 
dissolving  the  agar,  since  this  changes  the  agar  itself  into  other  car- 
bohydrates assimilable  by  fungi.  Although  parallel  cultures  were 
commonly  made  with  agar,  the  studies  of  metabolism  recorded  in  this 
paper  were  made  in  tube  culture  of  the  fluid  nutrients  only,  to  avoid 
possibilities  of  error. 

VARIOUS  SOURCES  OF  CARBON. 

Cane  sugar  (Tables  4.  and  5).—C&ne  sugar  was  added  to  Dox's  fluid 
in  the  following  percentages:  1.5,  3,  10,  20,  60,  and  75.  Of  the  spe- 
cies used,  but  one— P.  digitatum  of  Saccardo— failed  to  grow  typ- 
ically.    This,  together  with  other  work,  indicates  that  this  species  is 


84  CULTURAL-  STUDIES  OF   SPECIES  OF   PENICILLIUM. 

incapable  of  assimilating  nitrogen  from  the  sodium  nitrate  of  this 
solution.  Four  more  forms — P.  hrevicaule  and  its  closely  related 
varieties,  and  P.  roseum  Link — reached  normal  appearance  slowly. 
Other  cultures  indicate  that  these  forms  assimilate  nitrogen  in  this 
form  less  readily  than  in  organic  combinations.  This  medium  con- 
taining cane  sugar  in  amounts  as  great  as  20  per  cent  proved  well 
adapted  for  all  other  species  tried.  The  medium  as  used  was  neutral 
or  slightly  alkaline  in  reaction.  Thirty-three  of  the  forms  inoculated 
into  it  produced  pronounced  acid  reactions  to  litmus  in  a  few  days,  i.e., 
were  able  to  ferment  this  form  of  sugar.  In  prolonged  culture  with 
smaller  amounts  of  sugar,  some  of  these  forms  finally  reduced .  the 
acidity  and  even  brought  about  an  alkaline  reaction  again;  others 
remained  acid  as  long  as  observed.  Twelve  forms  failed  to  produce 
acidity.  This  number  includes  the  four  forms  in  which  growth  was 
delayed  and  P.  italicum,  P.  luteum,  P.  purpurogenum,  P.  roqueforti, 
and  P.  duclauxi. 

In  cultures  at  a  concentration  of  60  per  cent  cane  sugar,  five  forms 
produced  typical  colonies  at  once;  six  others  slowly  reached  normal 
proportions ;  a  few  more  grew  fairly  well ;  but  fully  half  the  species  tried 
produced  germination  with  but  little  further  growth.  Water  in 
amount  approximately  to  reduce  the  concentration  to  35  per  cent  was 
added  to  the  cultures  that  failed  to  produce  normal  colonies,  and  this 
was  followed  by  the  prompt  recovery  of  several  species  which  quickly 
reached  normal  development.  Critical  examination  of  the  data 
obtained  showed  that  closely  related  types  responded  in  exactly  the 
same  manner  to  changed  percentages  of  cane  sugar.  As  a  means  of 
separating  closely  related  forms,  further  determination  seemed  fruit- 
less. Exact  determination  of  the  maximum  percentages  of  cane 
sugar  tolerated  by  particular  species  has  not  been  completed.  The 
inhibition  is  not  a  stoppage  of  all  development  at  a  definite  critical 
concentration,  but  rather  a  gradual  reduction  of  activities  with  the 
increasing  concentration  of  the  medium.  Specific  maxima  and 
minima  are  therefore  almost  impossible  to  define.  All  determina- 
tions would  therefore  rest  upon  the  judgment  of  the  observer  rather 
than  upon  fixed  standards. 

Lactose  (Tahles  4  cif^  5). — Lactose  was  added  to  Dox's  fluid  in 
percentages  up  to  10  per  cent.  Prompt  and  normal  development  was 
determined  in  eight  forms ;  nine  more  forms  reached  typical  appearance 
more  slowly.  Only  twelve  of  these  produced  definite  acid  reactions  to 
litmus.  Among  the  forms  included  in  the  seventeen,  four  groups  of 
closely  related  organisms  were  found,  namely,  the  camemherti  group, 
Nos.  5, 6, 39 ;  the  common  green  group,  Nos.  22, 23, 40 ;  the  chrysogenum 
group,  Nos.  25,  26,  35,  and  44;  and  the  hrevicaule  group,  Nos.  2,  3, 
and  4.     With  lactose  as  with  cane  sugar  closely  related  forms  give 


COMPARATIVE  CULTURAL  DATA.  86 

approximately  the  same  reaction  in  most  cases.  Three  of  these 
groups  show  acid  and  the  fourth  alkahne  tests  with  htmus.  A  few  of 
the  remaining  species  continued  to  grow  until  after  several  weeks 
they  reached  almost  normal  development.  In  such  cases  the  assimi- 
lation of  carbon  from  lactose  seems  to  be  very  slowly  and  with  diflB- 
culty  accomplished  by  a  large  proportion  of  the  species  studied,  and 
to  be  practically  impossible  to  some  species. 

Comparison  of  the  litmus  reactions  produced  by  the  various  species 
with  cane  sugar  and  with  lactose  accounts  for  striking  differences  in 
this  reaction  when  litmus  is  introduced  into  gelatin  or  agar  media 
containing  these  two  forms  of  sugar.  A  species  which  will  ferment 
cane  sugar  and  not  ferment  lactose  will  produce  an  acid  reaction  with 
one  and  an  alkahne  reaction  in  media  containing  the  other. 

Lactic  add  {Tables  4  ^^  ^)- — Tubes  were  prepared  containing 
Dox's  fluid  to  which  0.9  per  cent  of  lactic  acid  was  added.  Thirty 
forms  were  inoculated  into  this  medium.  Of  these  forms  nine  pro- 
duced normal  and  typical  colonies,  showing  but  shght  inhibiting 
effect  from  the  acid.  As  many  more  cultures  slowly  became  typical 
colonies.  Nearly  every  form  germinated  and  produced  slight  growth. 
In  the  camemherti  group  (Nos.  5,  6,  39)  and  some  others  the  litmus 
reaction  became  alkaline.  It  was  thus  shown  that  a  series  of  species 
could  secure  carbon  from  lactic  acid  and  in  doing  so  destroyed  the  acid 
character  of  the  medium.  The  species  which  grew  most  rapidly 
in  lactic  acid  were  those  which  had  developed  best  in  the  lactose 
solutions. 

Levulose  (Tables  4  and  5). — Tubes  were  prepared  into  which  2.5  per 
cent  of  levulose  was  introduced  as  the  source  of  carbon.  The  re- 
sults as  tabulated  may  be  seen  to  group  together  about  the  same 
species  as  the  previous  experiments,  except  that  one  or  two  forms 
were  found  to  grow  well  in  levulose  that  failed  to  grow  with  lactose  or 
lactic  acid  as  a  source  of  carbon.  Fourteen  forms  produced  typical 
colonies  without  inhibiting  effects,  while  five  more  slowly  reached 
typical  development. 

Galactose  (Tables  4-  (^nd  5). — A  series  of  cultures  was  made  in  the 
same  way  with  3  per  cent  galactose  as  a  source  of  carbon.  Galactose 
proved  much  better  adapted  to  supply  carbon  than  either  lactose  or 
levulose.  Twenty-five  forms  produced  typical  growth,  and  others 
grew  more  or  less  readily.  This  form  of  sugar  therefore  proved  of 
but  small  assistance  in  the  separation  of  species. 

Glycerin  ( Tables  4  (ind  5) . — Cultures  offering  carbon  in  the  form  of 
3  per  cent  glycerm  produced  much  less  growth  than  those  containing 
sugars.  Eleven  forms  eventually  reached  fairly  typical  growth;  four 
only  of  these  showed  no  restraining  effect  of  the  medium.  Of  these, 
three  are  probably  closely  related  if  not  merely  races  of  a  single 
species — the  one  most  common  in  general  cultural  work  in  this 


86  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

region.  Apparently  glycerin  presents  a  form  of  carbon  much  less 
available  for  assimilation  by  species  of  this  genus  than  the  sugars. 

Butter/at  (Table  4)- — To  test  the  ability  of  these  fungi  to  assimilate 
fat,  butter  was  melted,  strained,  filtered  through  filter  paper,  and 
added  to  Dox's  fluid.  Although  not  chemically  pure,  perhaps,  it 
is  believed  that  the  amounts  of  other  nutrients  would  be  too  small 
to  affect  results.  Only  one  form  (P.  digitatum)  failed  to  grow. 
P.  luteum  gave  only  slight  growth.  The  majority  of  forms,  although 
growing  slowly,  produced  typical  or  fairly  characteristic  colonies. 
The  masses  of  fat  were  visibly  much  changed,  becoming  incrusted 
with  a  white  substance  in  most  cultures.  In  a  few  cultures  the 
action  of  fungus  caused  the  separation  of  the  various  fats,  so  that 
drops  of  yellow  oil  separated  out  from  the  remaining  nonliquid 
matter. 

Potato  starch. — In  one  series  twenty-seven  forms  were  cultivated 
in  Dox's  fluid,  containing  3  per  cent  of  potato  starch.  All  of  the 
common  species  of  the  genus  were  found  to  grow  normally  upon  a 
medium  containing  starch  as  the  source  of  carbon.  The  characters 
in  this  medium  were  approximately  the  same  as  in  the  stock  agar 
or  gelatin  cultures.  Two  species  (P.  decumbens  and  P.  digitatum). 
which  failed  to  grow  well  have  since  been  shown  to  depend  upon 
the  presence  of  cane  sugar  for  vigorous  growth  and  green  color  to 
their  spores.  Similarly  the  same  species  grown  upon  plugs  of  potato 
failed  to  produce  strong  colonies  of  pronounced  green  color. 

Malic  and  succinic  acids. — Series  of  cultures  were  made  with 
1  per  cent  malic  acid  and  with  1  per  cent  succinic  acid  as  sources 
of  carbon.  All  species  germinated,  but  no  species  reached  fully 
typical  development  in  either  series.  Some  few  species  produced 
slowly  colonies  of  half  or  more  of  the  normal  size  with  conidial 
masses  of  typical  color.  Man}^  of  the  species  grew  sufficiently  to 
produce  a  few  conidial  fructifications  recognizable  with  the  hand 
lens.  These  two  series  emphasize  the  observation  already  made 
that  species  inoculated  into  a  medium  ill  adapted  to  their  nourish- 
ment will  nevertheless  grow  and  produce  small  amounts  of  fruit 
under  widely  different  conditions  even  where  normal  growth  is 
impossible. 

CULTURES    IN    RAULIN's    FLUID    AND    COHN's    SOLUTION. 

The  comparative  data  for  cultures  in  Raulin's  fluid  and  in  Cohn's 
solution  are  shown  in  Tables  3  and  5.  Raulin's  fluid,  as  given  by 
Smith,  is  a  higldy  acid  medium  and  has  been  found  very  well  adapted 
for  the  growth  of  certain  species.  The  solution  is,  however,  too 
complex  to  make  analysis  of  cultural  results  upon  it  readily  possible. 
It  contains  carbon  in  three  different  forms — tartaric  acid,  potassium 
carbonate,  and  magnesium  carbonate — and  nitrogen  in  two  forms 


COMPARATIVE   CULTURAL   DATA.  87 

in  ammonium  nitrate.  Aside  from  data  as  to  growth  or  failure  to 
grow,  culture  in  such  a  medium  is  fioiitless. 

Cohn's  solution  is  also  an  acid  medium,  but  its  acidity  is  due  to 
the  presence  of  monopotassium  phosphate  (KIIjPO^).  In  a  series 
of  cultures  with  Dox's  formula  it  was  shown  that  the  monopotassium 
phosphate  had  practically  no  different  effect  upon  cultures  of  these 
species  than  the  dipotassium  phosphate  (KJiFO^). 

Very  few  of  the  species  were  found  to  grow  typically  upon  Cohn's 
solution,  however,  although  nearly  all  germinated.  Comparative 
work  with  Dox's  fornmla  shows  that  but  few  of  the  species  studied 
are  capable  of  normal  assimilation  of  carbon  from  forms  of  tartaric 
acid.  Only  three  of  the  species  experimented  with  failed  to  show 
distinct  germination.  One  of  these  inoculated  with  P.  camemberti 
var.  rogeri,  remained  four  months  without  exhibiting  any  germination 
of  the  conidia.  The  conidia  were  then  transferred  with  a  platinum 
needle  to  a  petri  dish  of  gelatin;  under  these  conditions  the  same 
spores  developed  into  typical  colonies  without  showing  any  ill 
effects  of  four  months'  immersion  in  the  fluid  which  they  were  unable 
to  assimilate. 

In  solutions  nontoxic  in  character  many  species  exhibit  definite 
selective  preferences  for  nutrient  elements  in  particular  chemical 
combinations.  This  selective  adaptability  to  particular  forms  of 
nutrients  differs  greatly  for  different  groups  of  species.  Some 
forms  of  wide  distribution  seem  adapted  to  produce  typical  growth 
upon  quite  varied  chemical  solutions.  Other  species  (e.  g.,  the  rots 
of  citrus  fruits),  equally  wide  in  their  distribution,  are  closely  depend- 
ent upon  particular  forms  of  food,  whereas  between  these  extremes 
are  many  forms  exhibiting  preferences  as  to  nutrients  yet  capable 
of  development,  although  more  slowly,  in  media  containing  nutrients 
in  forms  assimilated  with  difficulty.  These  experiments  have  offered 
no  tangible  evidence  of  rapid  adaptation  to  media  found  unadapted 
at  first  to  development.  Such  power  of  gradual  adaptation  is  not 
excluded,  however,  by  these  experiments. 

COLOR  IN  CONIDIAL  AREAS. 

In  dealing  with  all  species  it  has  been  found  by  repeated  experi- 
ment that  the  green  color  of  the  spores  is  dependent  upon  the  proper 
assimilation  of  the  carbon  element.  Many  of  these  species  produce 
a  brighter  green  when  cane  sugar  is  present  than  with  any  other 
form  tried.  Grown  upon  gelatin,  or  upon  potato  agar  or  bean  agar 
(free  from  sugar),  several  species  produce  green  masses  of  conidia 
which  rapidly  become  some  shade  of  gray,  brown,  or  almost  black 
in  media  without  sugar,  but  when  cane  sugar  is  present  they  are 
deep  green  and  remain  unchanged  for  much  longer  periods.  Certain 
other  species  entirely  lack  green  color  except  when  sugar  is  present. 


88  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

A  color  determination  is  therefore  dependent  for  its  value  upon 
knowledge  of  the  form  of  carbon  presented  by  the  medium.  These 
same  species  grown  with  lactose  as  a  source  of  carbon  respond  exactly 
as  they  do  in  potato  or  bean  agar  or  in  gelatin  free  from  sugar. 
Another  series  of  forms  are  little  if  at  all  changed  in  color  by  changing 
the  form  of  carbon  presented,  provided  only  that  they  are  able  to 
grow  readily  in  the  medium  as  presented.  It  seems  therefore  certain 
that  very  many  widely  distributed  species  are  capable  of  assimilating 
carbon  in  very  widely  different  chemical  forms  and  to  produce  in 
such  cases  normal  and  typical  growth  and  colors,  whereas  other 
species  are  entirely  dependent  for  normal  growth  and  color  upon 
the  presence  of  particular  chemical  combinations  or  groups  of  com- 
binations. Study  of  the  forms  so  responding  likewise  shows  that 
the  ubiquitous  species  are  capable  of  assimilating  carbon  in  the  most 
varied  combinations,  whereas  the  forms  lacking  this  power  are 
mostly  less  common  and  more  specialized. 

EFFECT   OF    CONCENTRATED    MEDIA. 

Similarly  great  differences  in  the  mass  of  the  growth  produced 
are  directly  attributable  to  the  presence  of  nutrients  easily  available 
or  in  greater  concentration.  The  formulae  usually  recommended  con- 
tain the  nutrient  used,  in  extremely  dilute  proportions.  The  deter- 
minations already  given  with  different  percentages  of  cane  sugar 
show  that  these  species  are  able  to  assimilate  sugar  in  widely  different 
concentrations.  With  the  majority  of  forms  studied  there  seems  to 
be  no  deleterious  effect  from  increasing  the  concentration  of  the  nutri- 
ents offered  until  the  solution  has  attained  an  osmotic  pressure  suffi- 
cient to  inhibit  growth  by  plasmolyzing  the  cells,  or  until  the  reduced 
percentage  of  water  gradually  reduced  the  rate  of  fungous  growth.  In 
one  experiment  nine  species  were  inoculated  into  a  medium  contain- 
ing 5  per  cent  cane  sugar,  10  per  cent  Witte's  peptone,  and  5  per  cent 
Liebig's  extract.  All  species  grew  luxuriously.  The  mass  of  myce- 
lium produced  was  much  in  excess  of  the  results  with  ordinary  culture 
media.  One  form  normally  producing  sclerotia  produced  a  very  rich 
growth  of  mycelium,  but  no  sclerotia.  In  media  of  higher  concen- 
trations the  amount  of  growth — the  mass  of  mycelium  and  conidia — 
is,  however,  so  greatly  increased  as  to  render  the  discrimination  of 
the  character  of  the  species  more  difficult  by  mechanical  interference 
due  to  the  quantity  of  material.  For  purposes  of  study,  therefore, 
the  usual  formulae  really  produce  the  more  satisfactory  growth  in 
nearly  every  species,  although  from  the  standpoint  of  fungus  devel- 
opment such  media  must  be  recognized  as  far  below  the  optimum 
concentration  for  the  species  of  this  genus. 

It  must  further  be  noted  that  in  cultures  containing  cane  sugar 
many  species  continue  to  produce  conidia  for  a  much  longer  period 


COMPARATIVE  CULTURAL  DATA.  89 

than  in  solutions  lacking  sugar  or  some  nutrient  equally  assimilable. 
In  these  cultures  the  quantity  of  conidia  produced  increases  enor- 
mously, often  becoming  a  layer  over  the  whole  surface  of  the  colony 
half  a  millimeter  or  more  in  thickness.  The  presence  of  such  masses 
of  spores  greatly  complicates  the  study  of  the  structure  of  the  colo- 
nies, but  is  especially  characteristic  of  such  species. 

THE   GROUPING   OF   SPECIES. 

Analysis  of  the  cultural  tables  presented  in  the  light  of  many  series 
of  comparative  cultures  makes  possible  the  grouping  together  of  par- 
ticular races  or  species  which  possess  common  cultural  characters. 
It  is  comparatively  a  simple  matter  to  single  out  first  the  unique 
forms — those  which  are  never  green,  those  which  produce  a  particular 
form  of  sclerotia,  those  which  regularly  produce  prominent  coremia, 
or  those  which  produce  striking  colors  in  the  substrata,  even  those 
associated  with  particular  substrata.  There  remain,  however,  the 
large  number  of  green  forms  which  lack  these  striking  characters. 
This  large  group  comprises  probably  most  of  the  forms  which  have 
masqueraded  under  the  name  P.  glaucum.  Lines  of  differentiation 
among  these  forms  are  more  or  less  obscure.  Kept  in  continuous 
culture  races  are  easily  differentiated  with  the  eye  by  shades  of  color 
or  habit,  but  characters  of  easily  recognizable  diagnostic  value  in 
written  descriptions  are  more  difficult  to  find.  Inspection  of  cul- 
tural data  shows,  however,  that  there  is  a  well-marked  group  of  these 
forms  which  are  able  to  ferment  lactose  as  well  as  cane  sugar.  These 
comprise  the  camemberti  group  (Nos.  5,  6,  39),  the  chrysogenum 
group  (Nos.  25,  26,  35,  44),  and  what  we  may  call  the  ''commune" 
group  (Nos.  22,  23,  40).  Among  those  not  causing  an  acid  reaction  in 
lactose  cultures  is  a  series  of  rapid  Hquefiers  of  gelatin  which  have 
many  characters  in  common  (Nos.  12,  15,  24,  37,  38).  All  of  these 
forms  show  special  adaptability  to  growth  in  cane-sugar  media.  No. 
15,  P.  citrinum,  produces  brilliant  lemon  color,  especially  in  sugar 
media;  No.  38  is  given  as  P.  atramentosum,  from  its  blackening  of 
the  substrata  in  sugar  media  and  in  milk ;  No.  37  by  developing  green 
color  when  sugar  is  present,  which  color  is  lacking  or  evanescent  with- 
out sugar.  These  forms  differ  in  their  color  reactions  in  the  medium, 
in  the  size  and  shape  of  their  conidia,  in  the  length  and  origins  of  their 
conidiophores,  and  in  the  arrangement  of  the  elements  in  the  conidial 
fructifications. 

Another  marked  habit  difference  which  holds  true  throughout  many 
series  of  cultures  is  the  tendency  of  colonies  of  certain  species  to 
spread  rapidly  over  the  whole  surface  of  the  substratimi,  whereas 
others  are  quite  restricted  in  their  habit  of  growth.  In  the  first  the 
developing  margin  is  almost  uniformly  broad  and  while  growing 
white,  e.  g.,  P.  rogueforti,  P.  italicum,  P.  chrysogenum,  P.  divaricaium, 


90  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

P.  spinulosum,  P.  expansum,  P.  hrevicaule,  and  such  floccose  forms  as 
P.  camemherti  and  P.  hiforme;  the  species  of  restricted  habit  with 
narrow  growing  border  are  represented  by  P.  citrinum  and  P.  atra- 
mentosum  and  their  allies.  The  designation  "spreading,"  "restricted," 
"with  broad  margin,"  or  "with  narrow  margin"  is  descriptive  for 
colonies  of  such  species. 

ODORS. 

Although  many  persons  seem  to  detect  the  presence  of  mold  by  its 
odor,  very  many  of  the  species  give  but  little  definite  odor.  A  few 
forms,  some  of  them  (e.  g.,  P.  hiforme)  always,  others  upon  special 
media,  produce  difinite  odors  by  which  they  can  be  recognized  or 
placed  in  particular  groups.  Grown  upon  gelatin  media,  P.  hrevi- 
caule produces  a  strong  ammoniacal  odor.  A  piece  of  moistened  Ut- 
mus  paper  held  over  such  a  colony  will  promptly  give  the  alkaline  re- 
action. This  organism  is  recorded  as  emitting  arsin  from  cultures  con- 
taining arsenic  in  any  form  and  it  is  said  to  be  a  very  delicate  test  for 
the  presence  of  that  element "  (Gosio) .  The  two  forms  here  described 
as  varieties  of  P.  hrevicaule  give  exactly  the  same  odor.  In  nearly 
every  medium  used  P.  expansum  (the  apple  rot)  gives  a  strong  odor, 
which  suggests  decaying  fruit  to  some.  Once  well  distinguished,  the 
presence  of  this  organism  can  be  detected  even  as  a  contamination 
wherever  it  occurs,  by  the  odor  alone.  Another  given  here  as  P. 
atramentosum  produces  a  very  characteristic  odor  while  digesting  milk, 
defined  by  one  as  the  odor  of  rancid  walnuts,  to  another  it  suggested 
mice.  A  series  of  forms  when  grown  upon  cane  sugar  produce  a  very 
characteristic  odor — an  ester,  according  to  the  chemists  to  whom  it 
was  submitted — recognizable  to  the  sense  of  smell,  but  not  definable. 
The  olive-colored  orange  rot  (P.  digitatum)  gives  this  most  strongly,  but 
it  is  also  given  by  P.  italicum,  P.  decumhens,  and  No.  13.  The  odor 
of  P.  claviforme  is  found  under  nearly  all  cultural  conditions,  and 
would  readily  identify  it  were  its  big  coremia  not  already  very  distinc- 
tive. The  common  and  undefinable  green  group  contains  a  series  of 
races  or  forms,  many  of  which  give  what  is  popularly  called  the  smell 
of  mold.     In  others  of  this  group  this  odor  is  scarcely  distinguishable. 

ANAEROBIC    CULTURES    (wiTH   CARBON   DIOXID). 

The  possibility  of  some  species  developing  under  anaerobic  con- 
ditions was  tested  as  follows:  Vials  were  prepared  with  Dox's  fluid 
having  5  per  cent  cane  sugar  as  a  source  of  carbon.  This  had  already 
been  shown  to  be  an  excellent  medium  for  the  growth  of  nearly  all  the 
species  used.  These  vials,  containing  all  the  species  herein  described, 
were  packed  in  a  crate  or  test-tube  basket  and  put  into  a  Novy  jar. 
The  jar  was  then  given  in  charge  of  Mr.  Dox.     The  air  was  exhausted 

o  Gosio.  Azione  di  alcune  muffe  sui  composti  fissi  d'araenico.  Rivista  d'Igiene 
e  Sanita  Pubblica,  Rome,  1892.    See  page  201. 


INCUBATION  EXPERIMENTS.  91 

as  completely  as  possible.  Carbon  dioxid  washed  through  water  and 
through  sulphuric  acid  was  carefully  introduced.  The  exhaustion 
was  repeated  and  the  gas  introduced  a  second  and  a  third  time.  The 
cultures  were  then  permitted  to  stand  one  week  and  examined.  A  cul- 
ture of  Oidium  lactis  among  the  species  of  PenidUium  was  found  to 
have  grown  some.  No  species  of  Penidllium  had  produced  a  colony. 
The  carbon  dioxid  was  then  three  times  exhausted  and  replaced  by 
air  and  the  jar  permitted  to  stand  a  second  week.  Normal  colonies 
of  every  species  were  produced.  Clearly,  therefore,  no  one  of  the 
species  of  Penidllium  under  experiment  was  capable  of  growing  in  an 
atmosphere  of  carbon  dioxid,  and  no  species  was  killed  by  exposure 
to  such  atmosphere. 

INCUBATION   TESTS. 

The  importance  of  temperature  in  determining  the  distribution  of 
fungi  in  nature,  and  in  controlling  their  presence  in  the  household,  the 
dairy,  and  the  storage  room,  made  the  determination  of  the  limits  of 
temperature  for  the  growth  of  these  species  desirable.  The  following 
incubation  experiments  were  therefore  made.     (See  Table  6.) 

1 .  Incubation  at  20  °  C.  This  was  repeated  several  times  with  fully 
checked  records. 

2.  Incubation  at  37°  C.  (range  of  variation  35°  to  38°  C.)  for  six 
days;  cultures  then  examined,  recorded,  and  the  incubator  cooled  to 
20°  C.  for  the  succeeding  six  days. 

3.  Use  of  the  ice  thermostat."  Four  series  of  cultures  were  made 
in  bean-agar  with  5  per  cent  cane  sugar — a  medium  adapted  to  pro- 
duce typical  colonies  of  all  species  in  a  minimum  of  time.  The  tem- 
perature of  the  incubator  was  recorded  eight  times  a  day  at  three- 
hour  intervals.  The  range  of  temperatures  in  the  compartments 
used  and  the  average  of  fifty-five  observations  taken  in  the  first  seven 
days  were  as  follows: 

Compartment  1,  average  1.05°  C,  range  0.5°  to  2°  C. 
Compartment  2,  average  4°  C,  range  3.2°  to  6°  C. 
Compartment  3,  average  7°  C,  range  6°  to  10°  C. 
Compartment  5,  average  8.7°  C,  range  7°  to  10.5°  C. 

Observations  upon  the  cultures  were  made  by  the  writer  at  3  and 
7  days  and  repeated,  by  the  kindness  of  Miss  Lucia  McCullough, 
of  the  Bureau  of  Plant  Industry,  at  15,  23,  and  29  days.  For  con- 
venience of  comparison,  the  notes  from  all  these  observations  have 
been  reduced  as  fairly  as  possible  to  a  decimal  code,  in  which  ger- 
mination of  conidia  without  further  growth  is  given  as  0. 1  and  typical 
colonies  at  1.0;  fractions  from  0.1  to  0.7  represent  vegetative 
mycelium  without  colored  conidial  areas,  and  from  0.7  to  1.0  the 

oBy  the  courtesy  of  Dr.  Erwin  F.  Smith  the  ice  thermostat  of  the  Bureau  of  Plant 
Industry  of  the  United  States  Department  of  Agriculture  was  placed  at  my  disposal. 
It  was  iced  and  regulated  under  his  instructions. 


92  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

completion  of  the  typical  colony.  The  figures  are  brought  together 
in  Table  6. 

Data  at  20°  C. — The  data  at  20°  C,  as  given  in  the  first  column  of 
Table  6,  are  regarded  as  ty])ical  for  the  species  studied  and  given  as  1.0. 
Numerous  series  of  cultures  with  all  these  forms  under  close  observa- 
tion in  the  incubator  at  20°  C.  and  in  the  laboratory  where  the  tem- 
peratures used  ranged  from  15°  to  25°  C.  or  slightly  higher  have  given 
approximately  the  same  results.  Within  these  limits,  rise  or  fall  in 
temperature  affects  the  amount  of  growth  or  the  stage  of  develop- 
ment of  the  colonies  within  a  specified  time  without  affecting  the 
character  of  such  growth.  The  differences  between  cultures  grown  at 
different  temperatures  within  these  general  limits  are  quantitative, 
not  qualitative.  Unless  made  for  a  specific  purpose  cultures  of  these 
fungi  may  be  safely  grown  outside  the  incubator  without  affecting 
their  character,  since  the  conditions  in  the  ordinary  working  room  are 
approximately  those  furnished  to  these  forms  by  nature. 

Data  at  37°  C. — At  37°  C.  thirteen  forms  showed  normal  develop- 
ment. Of  these  seven  grew  better  at  37°  C.  than  at  20°  C,  this 
number  including  but  one  well-known  species— P.  luteum.  At  the 
same  temperature  the  spores  of  seven  species  were  killed,  including 
among  these  P.  italicum  and  P.  digitatum,  the  species  destructive  of 
citrus  fruits.  Of  the  green  forms  abundantly  found,  only  one  grew 
well  at  37°  C. — P.  chrijsogenum.  The  numerous  green  forms  studied 
were  not  killed,  but  simply  prevented  from  growing  by  the  heat. 
Every  form  except  those  noted  as  killed  developed  normally  in  the 
same  tubes  as  soon  as  cooled  to  20°  C. 

Ice  thermostat. — In  compartment  1  of  the  ice  thermostat,  ranging 
from  0.5°  to  2°  C,  20  of  the  forms  experimented  with  either  produced 
germ  tubes  only  or  failed  even  to  germinate  in  twenty-nine  days.  Of 
the  remaining  18,  only  6  produced  colored  conidial  areas  in  that  time. 
Several  other  species  produced  considerable  masses  of  white  mycelium. 

In  compartment  2,  ranging  from  3.2°  to  6°  C,  with  an  average 
slightly  above  4°  C,  16  or  17  still  showed  germination  only  or  com- 
plete inhibition ;  1 1  showed  colored  conidia ;  several  additional  forms 
had  germinated  or  produced  distinguishable  mycelium. 

In  compartment  3,  with  an  average  temperature  about  7°  C.  and  a 
range  from  6°  to  10°  C,  16  forms  produced  colored  fruit.  Of  these, 
9  had  reached  the  typical  appearance  of  mature  colonies  of  the  species 
within  the  29  days.  Nine  only  remained  without  showing  some 
mycelial  growth  in  addition  to  germination. 

In  compartment  5,  averaging  about  9°  C.  and  ranging  from  7°  to 
10.5°  C,  all  species  had  germinated  and  all  but  one  had  produced 
mycelium;  25  forms  had  developed  colored  conidia;  1 7  had  produced 
colonies  of  typical  appearance. 


DISCUSSION   OF  INCUBATION   TESTS.  93 

Study  of  the  figures  given  shows  a  progressive  increase  in  growth 
from  the  coldest  to  the  warmest  compartment.  Examination  of  the 
cultures  showed  that  at  the  lower  limits  of  growth  very  many  species 
produce  colored  fruit  very  slowly  at  temperatures  at  which  vegetative 
mycehum  is  still  developed  quite  rapidly.  There  often  results, 
therefore,  in  cold  temperatures  a  disproportionate  growth  of  white 
mycelium  and  a  tardy  development  of  colored  conidia,  which  often 
affects  the  appearance  of  the  resulting  colony  considerably.  Cultures 
grown  under  such  conditions  would  be  diflicult  to  identify  in  many 
cases.  The  forms  which  grew  most  rapidly  at  37°  C.  either  failed  to 
grow  at  the  colder  temperatures  or  responded  very  slowly.  The  large 
majority  of  the  species,  and  especially  those  most  commonly  found  in 
food  materials,  are  seen  to  begin  fairly  rapid  growth  at  temperatures 
within  a  few  degrees  of  the  freezing  point.  When  taken  from  the  ice 
thermostat  all  cultures  which  had  failed  to  produce  normal  colonies 
grew  quickly  to  typical  appearance.  There  was,  therefore,  no  injury 
attributable  to  continuation  for  29  days  at  low  temperature.  The 
general  effect  of  low  temperatures  upon  these  species  is  the  suspension 
of  or  the  reduction  of  the  rate  of  development.  In  many  cultures 
the  beginnings  of  growth  were  diflficult  to  detect  and  in  most  cases  an 
exactly  critical  temperature  is  not  determinable,  since  cultures  not 
showing  any  growth  in  one  week  seem  gradually  to  adjust  them- 
selves to  conditions  and  produce  mycelium  in  the  succeeding  weeks. 

Eustace^  has  recorded  that  one  of  these  species  (P.  expansum 
Link  of  this  paper)  will  produce  rot  in  storage  apples  where  the 
temperature  of  the  room  as  recorded  does  not  rise  above  32°  F. 
(0°  C).  When  the  time  was  extended  to  two  months  Petri-dish 
cultures  under  the  same  conditions  produced  small  colonies.  The 
experiments  here  recorded  tend  to  suggest  that  very  little  growth 
will  occur  in  most  species  at  temperatures  nearer  than  2°  C.  to  the 
freezing  point,  although  many  of  them  will  germinate.  Unpub- 
lished records  of  the  temperatures  of  apples  in  storage,  furnished 
by  Mr.  C.  D.  Jarvis,  of  this  station,  showed  that  the  flesh  of  apples 
in  storage  was  constantly  from  1  to  2  degrees  at  least  above  that  of 
the  room.  Allowing  for  the  conductivity  of  the  thermometer  itself, 
the  difference  is  probably  somewhat  greater.  Both  series  of  data 
indicate,  however,  that  storage  temperature  to  exclude  fungous 
growth  must  be  close  to  the  freezing  point.  It  is  clear  that  low  tem- 
peratures (above  freezing)  merely  restrain  growth,  not  entirely 
prevent  it.  It  is  also  clear  that  by  restraining  the  production  of 
colored  fruit  many  colonies  would  be  rendered  inconspicuous  (al- 
though widely  growing),  thus  accounting  for  the  complaint  so  often 
heard  with  reference  to  dairy  products  taken  from  the  refrigerators, 
that  they  turn  green  with  mold  very  quickly. 


94  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

SUMMARY   OF   DATA   FROM    COMPARATIVE   CULTURE. 

1.  Species  closely  related  in  morphology  and  general  appearance 
give  closely  similar  reactions  in  culture  under  most  conditions,  but 
commonly  show  a  few  well-marked  differences  in  special  media  or 
under  special  conditions. 

2.  Certain  species  will  grow  in  media  of  widely  differing  composi- 
tion; others  require  particular  media  for  normal  development. 

3.  Cane  sugar  in  low  concentrations  is  readily  assimilated  by 
every  species  studied. 

4.  Butterfat  was  attacked  by  nearly  every  species. 

5.  Lactose,  galactose,  levulose,  and  glycerin  are  assimilated  by 
some  species  and  not  by  others. 

6.  Potato  starch  produced  normal  growth  in  most  species. 

7.  Very  few  species  grew  at  37°  C;  i.  e.,  very  few  species  could 
be  parasitic  to  warm-blooded  animals.  Few  species  were  killed  at 
37°  C,  and  species  not  killed  grew  normally  when  the  medium  was 
cooled  to  20°  C. 

8.  Incubation  at  low  temperatures  shows  that  growth  in  some 
species  will  begin  at  temperatures  very  close  to  the  freezing  point, 
but  that  only  a  few  species  will  actually  develop  in  cold-storage 
temperatures. 

9.  None  of  the  species  was  found  to  grow  in  an  atmosphere  of 
carbon  dioxid,  but  no  species  was  killed  by  such  atmosphere. 

10.  In  most  species  failing  to  grow  in  a  particular  medium  a 
change  of  concentration  or  the  addition  of  a  missing  element  will 
permit  normal  growth  unless  killed  by  osmotic  pressure  or  definitely 
toxic  agents. 

KEYS  TO  CULTURAL  IDENTIFICATION  OF  SPECIES. 

Media:  Prepare  the  following  media — 

1.  15  per  cent  gelatin '^  in  distilled  water. 

2.  15  per  cent  gelatin  in  distilled  water  plus  3  per  cent  cane  sugar. 

3.  Either  bean  or  potato  decoction  plus  1.5  per  cent  agar. 

4.  Bean  or  potato  agar  plus  3  per  cent  cane  sugar. 

Litmus  solution  may  be  added  if  desired  when  cultures  are  made. 

Prepare  Petri  dishes  with  10  c.  c.  of  each  of  the  media  used  and 
allow  them  to  cool.  Inoculate  tw^o  or  more  Petri  dishes  of  each 
medium  with  spores  of  the  species  under  examination.  Incubate 
at  20°  C.   (the  temperature  of  the  working  laboratory  is  usually 

oin  most  of  these  studies  the  "gold-label"  gelatin  imported  by  Bausch  &  Lomb 
from  Germany  has  been  used.  Culture  of  a  species  in  a  solution  of  gelatin  in  water 
has  two  uses  in  this  paper — the  detection  of  the  production  of  enzyms  capable  of 
liquefying  this  medium,  and  the  estimation  of  the  ability  of  the  species  to  grow  in  a 
medium  free  from  carbohydrates.  Neither  of  these  tests  seems  to  be  vitiated  by 
media  made  up  at  different  times  from  materials  of  different  origin.  Such  differences 
as  are  induced  by  the  differences  in  the  gelatin  are  quantitative,  not  qualitative. 


CULTURAL  IDENTIFICATION   OF   SPECIES.  95 

satisfactory).  Examine  at  intervals  of  three  days  or  less,  making 
observations  with  the  naked  eye  from  above  and  below,  with  the 
hand  lens  and  with  the  compound  microscope,  using  16  mm.  and 
8  mm.  or  6  mm.  objectives  to  determine  details  of  structure  and 
fruit  formation  from  growing  colonies.  A  drop  of  litmus  solution 
upon  the  margin  of  the  colony  will  test  acidity  or  alkalinity.  Ex- 
amine 1  and  2  for  liquefaction,  2  and  4  for  coremium  and  sclerotium 
formation.  Sclerotium  formation  will  be  found  to  call  for  continued 
examination  for  at  least  two  weeks. 

Two  separate  keys  are  presented  in  the  following  pages:  (1)  A  gen- 
eral key  to  all  the  forms  fliscussed  in  this  paper,  based  upon  cultures  in 
the  media  referred  to  above,  and  (2)  a  key  to  those  species  for  which 
presence  upon  a  particular  substratum  establishes  a  presumption  of 
identity. 

The  species  of  most  economic  importance  are  found  in  key  2. 
Key  1,  however  faulty,  is  an  endeavor  to  analyze  the  data  from 
many  series  of  comparative  cultures  in  such  a  way  as  to  simplify 
the  identification  of  the  forms  included  as  far  as  possible.  If  the 
complexity  of  the  data  offered  proves  a  barrier  to  identification  it 
may  be  hoped  that  it  will  also  deter  those  who  fail  to  identify  species 
clearly  from  using  specific  names  in  discussing  work  done  with  forms 
of  this  genus. 

Key  1. — Analysis  of  Species  in  Cultures  upon  Gelatin  and  Agar. 

A.  Species  fruiting  typically  by  coremia  (vertical  and  definite). 

a.  Coremia  long  (3-15  mm.). 

1.  Conidial  masses  strictly  terminal,  olive  green,  fragrant.  .P.  claviforme. 

2.  Upper  third  of  coremia  fertile,  conidia  green P.  duclatixi. 

aa.  Coremia  small: 

1.  Coremia  definite,  densely  crowded,  colony  orange  below, 

P.  granulatum. 

2.  Coremiform  character  indicated  in  cultures  by  clustering  of 

conidiophores,  definite  coremia  only  in  old  cultures,  becom- 
ing large  and  definite  upon  apples P.  expansum. 

AA.  Species  not  (or  rarely)  producing  coremia  in  culture. 

B.  Species  constantly  producing  sclerotia  or  ascigerous  masses. 

b.  Producing  ascigerous  masses,  yellow  or  reddish P.  luteum. 

bb.  Sclerotia  appearing  as  white  masses  in  old  cultures P.  italicum.<* 

bbb.  Sclerotia  reddish  or  pink,  globose  or  elliptical,  500;u  or  less  in  diameter. 

c.  Conidial  fructification  a  column: 

1.  Column  dense,  long,  sclerotia  partially  buried  in  substratum, 

P.  No.  30. 

2.  Column  formed  of  loose  chains,  scler  tia  numerous,  exposed, 

P.  No.  29. 
cc.  Conidial  fructification  of  divergent  chains: 

1.  Rapid  liquefier,  spores  globose,  2.5-3/1 P.  No.  31, 

2.  Slow  liquefier,  spores  elliptical,  3.5-4X2.5-3/i P.  No.  32. 

a  In  its  earlier  development  P.  italicum  Wehmer  will  be  usually  thrown  under  the 
head  kk,  on  account  of  its  habit  of  growth. 


96  CULTURAL  STUDIES   OF   SPECIES   OF   PENICILLIUM. 

BB.  Sclerotia  not  (or  rarely)  produced  (under  special  conditions). 
(Use  gelatin  cultures  (1)  and  (2),  compare  agar  cultures). 

C.  Rapid  liquefiers  (abundant  liquid  in  5  to  12  days). 

D.  With  definite,  strong  ammoniacal  odor: 

1.  Yellowish  brown-avellaneous  spores  rough P.  brevicaule. 

2.  White  or  cream,  spores  rough P.  brevicaule,  var.  album . 

3.  White  or  cream,  spores  smooth P.  brevicaule,  var.  glabrum. 

DD.  Without  ammoniacal  odor. 

E.  With  yellow  coloration  of  liquefied  gelatin  {not  of  mycelium  in  reverse). 

1.  Colonies  small,  conidiophores  100-150/f  in  length P.  citrinum, 

2.  Colonies  broadly  spreading,  conidiophores  250-300/i.  .P.  chrysogenum. 
EE.  Without  yellow  color  in  liquefied  gelatin  (or  slight  traces  only). 

e.  Colonies  white  to  pink  or  salmon P.  roseum. 

ee.  Colonies  some  shade  of  green. 

/.  Colonies  floccose,  margin  spreading  by  stolons P.  stoloniferum. 

ff.  Colonies  velvety-surface  growth  of  fruiting  hyphse  only. 
g:  Conidiophores  very  short  (100-200/t): 

1 P.  No.  12. 

2 P.  No.  37. 

gg.  Conidiophores  longer  (200-400;£): 

1.  Conidiophores  variously  branched,  reverse  always  colorless.  .P.  No.  24. 

2.  Conidiophores  each  with  a  verticil  of  branches — each  branch 

bearing  a  columnar  fructification — reverse  and  medium  dark- 
ened in  sugar  media P.  atramentosum. 

CC.  Liquefaction  of  gelatin  none  or  slower  than  10-12  days,  or  only  partial. 
G.  Colonies  never  green. 

h.  Colonies  yellowish  brown,  spores  elliptical P.  divaricatum. 

hh.  Colonies  white  to  lilac,  slow  liquefier,  14-16  days P.  liladnum. 

hhh.  Colonies  floccose  white  or  creamy: 

1.  Conidiophores  long,  typical  penicillate  branching, 

P.  camemberti,  var.  rogeri. 

2.  Conidial  chains  borne  upon  short  branches  of  floccose  hyphse, 

P.  No.  33. 
GG.  Colonies  some  shade  of  green.    . 

H.  Siu"face  with  hyphse  definitely  in  ropes  or  trailing,  bearing  numerous  conidio- 
phores as  short  branches  distinctly  traceable  to  their  origin  in  such 
hyphse. 
i.  Colonies  usually  red  below  and  reddening  the  substratum. 

1 .  Fruiting  areas  dark  green P.  funiculosum. 

2.  Fruiting  areas  mixed  yellow  and  green P.  pinophilum. 

ii.  Colonies  not  producing  red  color: 

1.  Coloniies  gray  rarely  greenish,  very  loose  floccose P.  intricatum. 

2.  Colonies  green,  conidial  chains  in  simple  compact  columns.  .P.  No.  28. 

3.  Colonies  gray  to  green,  hyphse  scattered,  creeping P.  decdmbens. 


CULTURAL   IDENTIFICATION    OF   SPECIES.  97 

HH.  Surface  hyphse  not  in  well-defined  ropes,  nor  trailing. 

j.  Surface  hyphse  woven  floccose,  course  of  hyphse  not  traceable. 

1.  Gray-green,  long  conidiophores,  no  odor P.  camemberti. 

2.  Gray-green,  shorter  conidiophores,  strong  odor P.  biforme. 

jj.  Surface  growth  at  margin  simple  conidiophores,  in  older  parts  both  floc- 
cose hyphse  and  conidiophores. 

1.  Gray -greenish,   branching  of  conidiophore  rather  loose,   odor 

none  or  slight P.  No.  22. 

2.  Green,  conidial  fructifications  rather  compact,  odor  definite, 

"moldy" P.  commune. 

N.  B. — There  are  probably  a  number  of  races  in  this  group. 
jjj.  Fruiting  surface  velvety — of  simple  conidiophores  or  conidiophores  borne 

so  close  to  surface  of  substratum  as  to  appear  simple: 
k.  Conidial  mass  a  dense  column  of  conidial  chains. 

1.  Column  from  a  single  verticil  of  basidia P.  spinulosum. 

2.  Column  from  a  verticil  of  brachlets  with  verticillate  cells  and 

chains P.  rubrum. 

kk.  Elements  of  conidial  fructification  not  in  a  column. 

I.  Conidia  smooth. 

1.  Green,  broadly  spreading,  ripe  conidia  globose,  4-5ju P.  roqueforti. 

2.  Green,  less  spreading,  conidia  elliptical,  medium,  commonly 

purpled P.  purpurogenum . 

3.  Gray  or  olive  green,  conidia,  5-7  by  3-5/i P.  digitatum. 

II.  Conidia  delicate  rugulose P.  rugulosum. 

Key  2. — Species  Determinable  from  Substrata. 

(In  these  species  the  substratum  establishes  a  presumption  of  identity.) 
Cheese  (Camembert  and  Brie): 

1.  Floccose,  white  unchangeable,  no  odor P.  camemherti  var.  rogeri. 

2.  Floccose,  white  to  gray-green,  no  odor P.  camemberti. 

3.  Powdery,  yellowish  white,  spores  smooth,  ammoniacal  odor, 

P.  brevicaule  var.  glabrum. 

4.  Powdery,  yellowish  white,  spores  tuberculate,  ammoniacal  odor, 

P.  brevicaule  var.  album. 

5.  Forming  yellowish-brown  areas,  spores  rough,  ammoniacal  odor.  .P.  brevicaule. 
Cheese  (Roquefort): 

1 .  Green  streaks  inside  the  cheese P.  roqueforti. 

Citrus  fruits: 

1 .  Colonies  of  mold,  blue-green P.  italicum. 

2.  Colonies  of  mold,  olive-green P.  digitatum=olivaceum. 

Pomaceous  fruits  (apples,  pears,  etc.): 

1 .  Blue-green  colonies  finally  producing  coremia P.  expansum. 

Polyporacese  (Boleti,  Polypori,  etc.): 

1.  Colonies  green  (yellowish  green)  spreading  by  stolons P.  stoloniferum. 

Wood  (pine): 

Producing  orange  to  red  stains  in  pine  wood P.  pinophilum. 

8108— Bull.  118—10 7 


98 


CULTURAL  STUDIES  OP  SPECIES  OP  PENICILLIUM. 


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100  CULTURAL  STUDIES   OF  SPECIES  OF  PENICILLIUM. 

Table  2. — Gelatin  and  color  reactions. 


Spiecies. 


Name. 


pinophilum. 
brevkauk... 


var.  glabrum . 

var.  album  . . , 

camemberti 


var.  rogert . . 

clavi/orme 

lilacinum 

granulatum . . . 


italkum . 
luteum.. 


expansum . 
dtrinum . . 


digitatum 

purpurogenum . 


roqueforti. 
roseum . . . 


dudauxi. 
rubrum . . 


commune 

cf.  2G '. 

chrysogenum.. 
stoloniferum . . 


divaricatum. 


cf.  26 

decumbens. 


38  atramentosum . 

39  biforme 

40  I 

41  =19 

42  I  funiculosum. . . 
43 

cf.  26 

spiniilosum. 


rugulosum. 
intricatum. 


=38. 


15  per  cent  gelatin  in  water. 


Liquefaction. 


Very    slow,    6 

weeks. 
Rapid,   5    to   6 

days. 

'.'.'.'.do'.'.'.'.'.'.'.'.'.'. 
None    or    very 
slow. 

do 

Slow 

14  to  16  days 

None 


Litmus. 


Acid 

Alkaline. 


.do. 
.do. 
.do. 


None  or  slow 

None 

6  days 


None  in  15  days. 
Partial,  14  days . 

Rapid,    4   to   6 

days. 
None  in  15  days. 
....do 


....do 

Rapid,  8  days. 


None  in  15  days. 

(?) 

Partial,    15    to 
20  days. 

do 

Rapid  StolOdays 
do 


.do. 


Rapid,    6   to   8 

days. 
Very  slow,  20+ 

days. 
Rapid 


Very  slow. 
Rapid 


Partial,   slow, 

15+  days. 
None 


...do. 


Rapid 

None  or  ? 

Rapid,   6   to   7 

days. 
Rapid,6tol0days 

None  in  15  days. 
do 

Rapid 

None 

None  or  ? 

Rapid 

Partial,  slow 

None  or  ? 


Rapid. 
None.. 


None  or  ?  slow. 

Partial  and 

slow. 
Rapid 


do... 

Neutral . . 
Alkaline. 
do... 


Faint  alkaline. 

Acid 

Alkaline 


.do. 
.do. 


.do. 


Alkaline . 
do... 


See  color 

(?) 
Slow  alkaline 


Alkaline. 

do... 

do... 


do 

do 

Acid 

Alkaline 


Acid 

Alkaline. 


Neutral?. 
Acid 


.do. 


Alkaline. 
Neutral?. 
Alkaline . 


do... 

do... 

do... 

do... 

Acid 

Neutral?. 
Alkaline . 
Acid 


Neutral?. 


Alkaline. 
.....do... 


.do. 
-do. 
.do. 


Color,  a 


In  conidial 
mass. 


Green. 


In  medium. 


Yellow  to  red.. 


Drab  to  choco-     None . 
late. 

White  or  cream do. 

do ..^ do. 

Gray-green do. 


White 

Olive 

White  or  lilac. 

Yellowish 

green. 


Bluish  green . . ;  None . 


Green,  small 
areas. 

Pale  blue- 
green. 

Deep  green 

Green  to  gray- 
green. 

Green 


do 

Brownish 

None 

Yellow    to 
orange. 


Reverse  of 
colony. 


Olive. 
Green. 


None  or  ? . 
None 


.do. 
.do. 


Lemon  yellow. 

None 

Red  or  no:ie... 


do !  None 

White  or  sal-   do 

mon. 

Olive  or  green. 

Green  or  vari- 
ous. 
Gray-green 


Green 

Deep  green. 
Green 


fYellow,  acid.. 

^Red,  alkaline.. 

(?) 

None 


.do. 


Yellowish 
green. 

Gray-green  to 
green.- 

White,  some 
green. 

Green 

Gray  or  green- 
ish. 

Gray-green 


White 

cream. 
Yellowish 

brown. 


or 


Gray  or  green. 
Gray  to  blue- 
green. 

Deep  green 

Gray-green 

Green 

(?) 

Green 

Light  olive 

Green 


.do. 


(?) 
Gray  to  green- 
ish. 
Yellowish 
green. 
(?) 

(?) 


do 

do 

Yellow  in  cer- 
tain media. 
None  or  golden 

None  or  ? 


Yellow  I  violet! 
None  or  slight. 


None 

None  or  yel- 
lowish. 

None  or  rod- 
dish. 

None 


.do. 


None. . 
do. 


None  or  brown 

None 

do 

(?) 
Red  or  none. . . 
Red 

None 


None  or  yel- 
lowish. 
(?) 
None 


.do. 


(?) 

(?) 


Red. 

None, 

Do. 

Do. 

Cream. 

Do. 
Brownish. 
None. 
Yellow      to 

orange. 
Brownish 

areas. 
Yellow      to 

orange. 
None. 

Do. 

None  or  brown- 
ish. 

None  or  yel- 
lowish. 

Brownish. 

Yellow  to  red 
or  none. 

None. 

White  or 
cream. 

Yellow,  acid. 

Red,  alkaline. 

Reddish. 


or 


White 
cream. 
Do. 
Do. 
White  or  yel- 
lowish. 
None    or    yel- 
lowish. 
Cream  or  yel- 
lowish. 
None  or  yellow- 
ish. 
Do. 


to 


None. 

None    or    yel- 
lowish. 

Yellowish 
salmon. 

Cream. 

None. 


White. 
Do. 

None  or  brown. 
Cream. 
Do. 
(?) 
Red  or  none. 
Orange  to  red. 

Cream  or  vio- 
let. 

Yellow  to 
orange  red. 

White  or  sul- 
phur. 
Uncolored. 


o  "'None"  means  the  absence  of  a  definite  color,  varying  from  hyaline  to  cream  at  times. 


COMPARATIVE   CULTURAL  DATA. 
Table  3. — Comparative  cultures. 


101 


No. 


Cohn's 
solution. 


Qenninated . 

Slow,  but 

typical. 
G  ermlnated 

only. 

do 

Fair,  white.. 

No  growth... 
Slight 


Weak,  typ- 
ical. 

G  erminated 
only. 

do 

do 

Slight 


Small  growth 
Weak 


Good,  no  yel- 
low. 
Germinated  . 

do 

Slight  growth 


do 

Germinated 


Fair  growth . 

Small  colo- 
nies. 

Slowly  typ- 
ical. 

Weak 

Not  typical. . 


Typical 

Weak  growth 


Germinated  . 

Slowly  typ- 
ical. 

Small  colo- 
nies. 

No  growth... 

Germinated  . 


Raulin's 
fluid. 


Germinated. 

do 

do 

Very  heavy. 

do 

Typical 


Typical 

do 

Slow,yellow. 
Slow,    typ- 
ical. 

Good 

Typical 


Good,  some 
yellow. 

Some  growth 

Slow  growth. 

Richly  typ- 
ical. 


Typical. 


Typical 

Slow,typical 


Typical 

Slowly  tyj)- 
ical. 

Typical 

do 


MUk  -i-CaCls, 
curdling. 


Slo  w  , 

days. 

10  days... 

13  days... 

8  days 

9  days 


15 


do 

do 


None. . . 
do. 


10  days.. 


Milk. 


Rate  of 
digestion. 


Very  slow. . 

Rapid 

do 


do 

Slow 


do... 

do... 


.do. 
.do. 


do... 


Coloration. 


Red... 
None. . 
do. 


...do... 
...do... 


do 

Reddish 

brown. 
None 


7  days. 

10  days 

8  days. . 


Very  slow  growth  in  milk. 


Slowly  red . 
Yellowish . . 


.do 


Rapid. 


13  days. 
8  days.. 


Slow do 

do Yellowish 

brown. 

Rapid Slight    yel- 
low. 
None;  poor  growth  in  milk 


None. 


Natural 
substratum. 


Wood  stain- 
ing. 
Soft  cheese . 

do 


do 

Camembert 

cheese. 
do 


Citrus  fruits. 


Apples. 


Rapid.. 
9  days. . 

13  days. 
8  days.. 

7  days. . 

8  days. . 


do. 

7  days. 


Germinated  . 

Weak  growth 
Germinated  . 

Half  typical. 
do 


Germinated  . 
No  growth... 


Germinated . 


Weak  growth 


Small    colo- 
nies. 


Not  typical. 
Good  growth 

Typical 

do 


do 

8  to  9  days . . 

9  davs 

do 

do 


Slow. 
Rapid. 


do. 

Slow.. 


.do. 
.do. 
.do. 


Rapid. 


.do. 
.do. 


Richly  tyi)- 
ical. 

Typical 

Weak  growth 


Typical. 


Typical. 


Very  slow. . . 

From    top 

down. 
Slow 


9  days 

Very  slow,  4 
weeks. 


9  days. 


8 davs 

do 

9  days 

Very  slow. . 


9  days. 
7  days. 


Very  slow. . . 

9  days 

8  days 

Slo w  ,     3 

weeks. 
None,    4 

weeks. 
11  days 


do. 

Slow.. 

do. 

do. 

Rapid. 

Slow.. 

Slight. 


Little  or 

none. 
Medium 

rapid. 
Little  or 

none. 
Fairly  rapid. 
Rapid 


Rather  slow. 

do 

do...... 

Little 


Slow,  slight 
Rapid 


Slow 

Slight 

Slow 

Very  slow. 


do. 

Slow.. 


Slight  red. 

None,  or  | 
evanescent 
greenish. 

None I 

Y  e  1 1  o  w  to 
red. 

None 

....do I 

....do ! 


Oranges. 


Roque  fort 
cheese. 


.do. 


Pale  yellow 

Golden  yel-  i 

low. 
Slight,  if  any    Fleshy  fungi. 


Pale  yellow. 

None 

do 

do 


do.. 

do.. 

do.. 

YeUow. 
None. . . 


do 

Reddish 
brown. 

None 

do 

do 

Red  where 
touched. 

Not  colored. 

Golden  yel- 
low. 

None 

do 

do 

do 


.do. 
.do. 


Dataat37»C. 


Grew  best. 

Grew  well. 

No  growth. 

Do. 
Do. 

Do. 
Do. 

Best  growth. 

No  growth. 

Killed. 
Best  growth. 
Killed. 

No  growth. 
Do. 

Slight 

growth. 
Killed. 
Grew  well. 

None. 


Killed. 
No  growth. 


Do. 
Do. 

Do. 

Slow  growth. 
Good  growth. 

Killed. 

Do. 

No  growth. 
Do. 

Grew  best. 

KiUed. 

Grew  best. 


Slight 

growth. 
KUIed. 
No  growth. 

Do. 
Do. 

Grew  well. 

Do. 

Grew  best. 
No  growth. 

Grew  best. 

No  growth. 


102  CULTURAL  STUDIES   OF   SPECIES  OF  PENICILLIUM. 

Table  4. — Comparative  cultures  in  synthetic  fluid  {Vox's). 


Carbon  supplied  i 


Cane  sugar. 


1  to  20  per  cent. 


Typical 

Slowly  typical 


do.. 

....do.. 
Typical . 


.do. 
.do. 

.do. 

.do. 
.do. 
.do. 
.do. 
.do. 
-do. 
.do. 


Not  typical . 


Typical 
do.. 


60  per  cent. 


75  per  cent. 


No  growth 

Germi  n  a  t  c  d     Slight  growth . . 
only. 

Slight ' do 

do do 

Slowly  typical..   Slow 


....do 

Nocoremiaijut 

acid. 
Slight 


Not  typical . . 


Typical . 

do.. 

do.- 


.do. 
.do. 
.do. 

.do. 


Typical 

Not  typical . 

Typical 

do 


.do. 


.do. 
.do. 


.do. 


.do. 
.do. 


do 

Slowly  typical 
Typical 

do 

do 

do 


.do. 

.do. 
.do. 
.do. 

.do. 
.do. 


Not  typical 

Slow,  typical... 

No  growth 

Good,  acid 

do 

....do 

Wealc,  no  yel- 
low. 

Germinated 
only. 

do 

Not  typical 

No  growth 

Germ  i  n  a  t  e  d 
only. 


Slowly  good 

do 

Weak 


Good.. 
....do. 
....do. 


.do. 


Green,      no 

sclerotia. 

....do 

Weak , 

Germ  i  n  a  t  e  d 

only. 
Slight 


Good 

Very  weak . 


Slowly  typical.. 


.do. 
.do. 


....do 

Weak 

do 

do 

Slowlytypical. 
do 


Fair,  not  typ- 
ical. 

Typical 

Weak 

Very  small  col- 
onies. 

Typical 

Slowlytypical. 


Slight. 


Lactose. 


No  growth 

Germinated 

No  growth 

Germinated 

Slow 

do 

Not  typical 

Weak,  hall  typ- 
ical. 
No  growth 


....do 

Weak 

No  growth . 
....do 


....do 

Very  pnwr 

do 

Germinated, 
small. 

Good 

do 

do 


Fair 

No  growth . 


Slight. 

No  growth . 
do 


.do. 


Good  growth... 
Pinhcad    colo- 
nies. 
Il all  typical 


Very  weak . 
ight 


Sligh 


do 

No  growth 

do 

do. 

Good  growth... 
Pinhead    colo- 
nies. 
Germinated 


Weak 

No  growth 

Pinhead    colo- 
nies. 

Good 

Weak 


Growth. 


Not  typical 

Slow,  typical... 


....do.. 

do.. 

Typical . 


do 

Very      weak 

growth. 
Germ  i  n  a  t  c  d 

only. 

Slow,  weak 

Slight 

do 

Weak,  typical. 
Slow,  typical., 
.do 


Litmus. 


Germinated do 


do 

Weak 

Slight 

Germinated. 


....do 

Rich  growth . . 
do 

Small  growth . 


Typical 

Slowlytypical. 
Weak 


Typical 

Slight 

Slowlytypical. 


Slight.. 

do. 

do. 


.do. 


Typical . 
Sfight... 


Slowly  growing 

Slowlytypical.. 
Typical 


do.. 

Slight... 

do.. 

do.. 

Typical . 
Sfight. . . 

do.. 


Good 

do 

Germinated. 


Slight 

Slow,  typical. 


No  effect 

do 


do. 

do. 

.\cid.. 


....do... 
No  effect. 


.do. 


....do... 

Acid 

No  effect. 

.do... 

.do... 

.do... 


Slow  growth..  .1 do 


0.9  per  cent  lac- 
tic acid. 


Germinated. 


Typical,   alka- 
line. 
Do. 
Germinated. 

Slight. 

Not  typical. 
Slight. 

Do. 
Weak. 
Good. 

Do. 
Slight. 

Germinated. 


.do Slow. 

.do Weak 

.do 1 

.do.... 


do... 

Acid 

do... 

No  effect. 


Good,  acid. 


Good. 
Do. 


.\cid . . 
....do 

No  effect \  Small 

nies 
.Vcid.... 
No  effect 
.\cid.... 


No  effect. 

do... 

do... 


.do. 


Acid 

No  effect. 


.do. 


do. 

Acid.. 


Typical. 

colo- 
Not  typical. 
Weak. 


No  growth. 
Good. 


Fair,  not  typ- 
ical. 

Do. 

Small    colo- 
nies. 

No  growth. 

Typical,   alka- 
line. 

do... 

No  effect. 

do Little  growth. 

do I         Do. 

Acid 

No  effect. 

do... 


Acid 

No  effect. 
do... 


.do. 
.do. 


COMPARATIVE   CULTURAL  DATA.  103 

Table  4. — Comparative  cultures  in  syntlietic  fluid  {Box's) — Continued. 


Carbon  supplied  as— 

No. 

2.5  per  cent  levu- 
lose. 

3  per  cent  galac-        3  per  cent  glyc- 
tose.                          erin. 

1 

Butterfat. 

Growth. 

Color  in  medium. 

1 

Good,  acid 

Fair,  alkaline 

Fair.acid 

Fair,  alkaline 

Slight 

Typical,  slow 

Yellow  to  red. 

? 

^ 

do... 

Fair 

4 

Slight  colony.. . 

do 

«i 

Typical 

Typical Slow,  typical 

do do     _ 

Slow,  typical 

do 

f, 

do.. 

Do. 

7 

Weak 

Good,  acid. .  .  . 

No  coremia 

do 

Tinged. 

Tinged  brownish. 

R 

Slow 

Slow,  alkaline 

Typical 

Fair,  acid 

Good,  acid 

Slight 

do 

9 

Slow,  small 

Not  normal 

Slight 

do 

do 

10 

No  growth . . 

do 

None. 

11 

Germinated 

Slow,  not  typical. . 
Goo  J 

Slight 

Do. 

1? 

Good,  alkaline 

Weak 

Do. 

n 

do do 

do do 

Small  colonies do 

Germinated Slight 

Small  colonies j  Small,  acid 

Weak 1  Good  colonies 

Slight 

Good 

Do. 

14 

Slow 

Small  colonies 

Germinated 

No  growth 

do 

Do. 

15 
16 

do 

None 

Lemon  yellow. 

17 

Slow 

Slowly  red. 

IS 

Good 

1<» 

Slight Slowlv  tvoical 

Do. 

?n 

Weak 

Weak 

do 

do 

Yellowish. 

?i 

n 

Good 

Typical 

Typical 

Typical 

None. 

r? 

.do 

do  . 

do 

do 

Do. 

?4 

Small 

.do 

Weak 

do 

Do. 

?.■) 

Typical 

do 

do 

?« 

?7 

Typical 

Typical 

Fair  colonies . . 

?8 

do 

Typical 

Reddish  brown. 

?9 

Slow 

None. 

30 
31 
32 

33 
34 
35 
36 
37 
38 

Fair.. 

Typical. . 

Very  weak 

.do 

Very  slow. 

Do. 

Typical 

do 

Very  slow,  typical. 
do 

Do. 

Slowly  typical 

do 

Very  small  colo- 
nies. 

Slowly  typical 

Germmated 

Do. 

Small  growth 

do 

Slowly  typical 

Weak 

Slightly  rusty. 
None. 

do 

Slow..  .. 

Small  colonies 

do.... 

Small  growth 

Tvrical.. 

Germinated 

Slight 

Weak 

Do. 

Slow.. 

Do. 

Slowly  typical 

do 

Germinated 

Typical 

Typical 

Do. 

39 

Typical •.. 

do 

do 

Do. 

40 

do 

do 

do 

do 

Do. 

41 

42 
43 

Little  growth 

...  .do 

Little  growth 

do 

Germinated 

do 

Coremiform   colo- 
nies. 
Slow,  typical 

Do. 
Yellowish. 

44 

45 

Typical 

Half  typical 

Slow 

Typical 

40 

Good 

Good 

Slowly  typical 

Slight  yellowish. 

47 

49 

Typical 

Typical 

Slow 

Fair 

51 

Half  typical 

do 

Slowly  typical 

Small,  typical T)  i 

104  CULTURAL  STUDIES   OF   SPECIES  OF   PENICILLIUM. 

Table  5. — Decimal  summary  of  comparative  cultures  in  synthetic  fluid  (Box's). 


No. 


Species. 


Name. 


r>inophilum 

hrevicaule 

var.  glabrum 
\a,T.  album. . . 

camemberti 

var.  rogeri 

claviforme 

lilacinum 

granulatum 

italicum 

luteum 


expanaum 

citrinum 

digitatum 

purpuTogenum . . 

roqueforli 

roseum 

dtKlauxi 

Tubrum 


commune 


chrysogenum. 
stoloniferum . 


divaricatum. 
cf.  26 

decumbens.. 


atramentosum 

biforme 


funiculoaum . 

cf.  17 

cf.26 

spinulosum.. 
rugulosum... 
iniricatum.. . 


Carbon  supplied  as— 


Cane  sugar. 


Up  to  20 
per  cent. 


1.0 
-1.0 
-1.0 
-1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 

(?) 

1.0 

1.0 

-1.0 

1.0 


1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 


1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 


1.0 
0.0 
0.0 
0.0 
1.0 
1.0 
1.0 

(?) 

1.0 
0.0 
0.0 
1.0 
1.0 
1.0 
1.0 

(?) 

0.0 
0.0 
0.0 

(?) 


1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 


1.0 
1.0 
1.0 

(?) 

1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 
1.0 


0.1 
0.1 
0.1 
0.1 
-1.0 
-1.0 
0.2 
0.1 
0.3 
-1.0 
0.1 
0.1 
1.0 
1.0 
0.5 
0.1 
0.1 
0.3 
0.0 
0.1 


-1.0 

-1.0 

0.4 

0.7 

0.7 


hi  o 


0.0 
0.1 
0.1 
0.1 
0.5 
0.5 
0.2 
0.0 
0.1 
0.0 
0.1 
0.6 
0.8 
0.4 
0.5 
0.0 
0.0 
0.2 
0.0 
0.0 


0.3 
0.6 
0.3 

0.7 
0.7 


1.0   -1.0 
1.0    -1.0 


0.0 

0.4 

0.5 

0.3 

0.1 

0.1 

1.0 

0.2 

0.5 

0.7 

-0.9 

-0.9 

0.1 

0.1 

0.7 

-0.8 

-0.6 

0.3 

-1.0 


0.2 


0.2 
0.0 
0.0 
0.0 
0.7 
0.5 
0.8 
0.3 
0.4 
0.4 
0.0 
0.0 
0.8 
0.5 
0.1 
0.0 
0.8 


Lactose. 


0.0 

-1.0 

-1.0 

-1.0 

1.0 

1.0 

0.2 

0.1 

0.5 

0.3 

0.2 

0.7 

-1.0 

-1.0 

-0.6 

0.1 

0.1 

0.5 

0.2 

0.1 


-1.0 
-1.0 
0.6 
1.0 
-1.0 
0.6 
1.0 


-1.0 
0.2 
0.2 
0.2 
0.2 
1.0 
0.2 
0.5 
0.9 
1.0 
1.0 
0.2 
0.2 
1.0 
0.2 
0.2 
1.0 
0.2 


1.0 
0.8 


1.0 
1.0 
0.4 
0.4 
0.5 
0.5 
0.2 
1.0 
1.0 
1.0 
0.6 
0.1 
0.3 
0.5 
0.2 
0.2 


1.0 
1.0 
0.6 
1.0 


1.0 
-1.0 


1.0 
1.0 
-1.0 
0.3 
0.3 


0.3 
0.5 
0.9 
1.0 
1.0 
0.2 
0.2 


1.0 
1.0 
1.0 


0.2 
0.1 
0.1 
0.3 
-1.0 
-1.0 
0.3 
0.2 
0.4 
0.3 
0.1 
0.4 
-1.0 
-1.0 
0.6 
0.1 
0.0 
0.4 
0.2 
0.2 


-1.0 
-1.0 
0.5 
0.7 
0.7 
0.9 
0.9 


0.2 
0.2 
0.3 
-1.0 
0.1 


0.1 

0.5 

0.1 

-1.0 

0.8 

0.1 

0.1 

0.5 

0.6 

-1.0 

-1.0 

0.1 


0.8 
0.8 


1.0 
1.0 
0.9 
0.8 
1.0 
0.9 
1.0 
1.0 
1.0 
1.0 
1.0 
0.4 
0.4 
0.8 


0.1 


0.3 


1.0 
1.0 
1.0 
1.0 


1.0 
1.0 
1.0 
1.0 
0.4 


0.4 
1.0 
1.0 
1.0 
1.0 
0.3 
0.3 


0.9 
1.0 
1.0 


1.0 
1.0 
0.1 
0.3 
0.6 
0.3 
0.3 
0.4 
0.8 


0.4 
0.1 
0.6 
0.4 


0.8 


1.0 
1.0 
1.0 


0.1 
0.1 
0.1 
+  1.0 
+  1.0 
1.0 


1.0 
1.0 
-0.7 
-1.0 
1.0 
1.0 
1.0 
0.4 
0.6 
1.0 


1.0 


1.0 
-1.0 


0.5 
0.5 


0.4 
0.0 
1.0 


0.7 


0.7 
0.7 
0.0 
1.0 


0.3 
0.3 


1.0 
-1.0 
1.0 
1.0 


0.5 
0.8 
1.0 
1.0 


1.0 
1.0 
0.5 


1.0 


1.0 


0.1 
-1.0 
0.1 
0.1 
0.8 
0.0 
0.2 
-0.7 
0.1 
0.1 
0.1 
0.2 
0.4 
0.5 
0.7 
0.1 
0.1 
0.2 
0.2 
0.1 


0.8 
0.5 
-1.0 
0.5 
0.5 
1.0 
0.6 


0.1 
-1.0 
0.6 
0.0 
0.1 


0.1 
0.3 
0.1 
0.7 
0.7 
0.1 
0.0 


0.1 


0.3 


(?) 


(?) 


(?) 


(?) 


Explanation  of  decimals:  1.0  denotes  normal  development,  or  present;  —1.0,  slowly  typical;  0.1,  ger- 
mination of  spores  only;  decimals  from  0.1  to  1.0  denote  estimated  amount  of  development  between  mere 
germination  and  t3rpical  development;  +1.0  denotes  very  rich  growth. 


COMPARATIVE   CULTURAL  DATA. 
Table  6. — Incubation  experiments. 


105 


20°  C. 

Ice  thermostat. 

Species. 

Six  days 
at  37'  C. 

! 

Compartment  1,'  Compartment  2, 
0.5-2°  C.        1 3.2-6*  C,  av.  4°  C. 

Compartment  3, 

e-W"  C,  av. 

7''+C. 

Compartments, 

7-10.5*  C,  av. 

8.7*  C. 

Growth ,  periods  Growth ,  periods 

Growth ,  periods    Growth ,  periods 

in  days. 

in  days. 

in  days.                 in  days. 

Name. 

2 

0 

2 

0 

i 

i 

7. 

15. 

23. 

29. 

7. 

15. 

23. 

29. 

7. 

15. 

23.. 

29.    7.  '  IS. 

1 

23. 

29. 

1 

vinophilum 

hrevicaule 

1.0 

+1.0 

0.0 

0.1 

0.1 

0.1 

0.1 

0.1 

0.1? 

0.1? 

0.1? 

0.1 

0.1? 

0.1? 

1      1 
0.1?  0.1 0.3 

0.5 

0.6 

2 

1.0 

1.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0. 0  0. 0 

0.1 

0.2 

0.3  0.40.5 

0.6 

J.  7 

3 

var.  glabrum. 

1.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.0 

0.2 

0.4 

0.4  10.3  0.4 

O.S 

0.7 

4 

var.  album. . 
camemberti 

1.0 
1.0 

0.0 
0.0 

0.0 
0.0 

.") 

0.20.5 

0.5 

0.6 

0.3 

0.5 

0.5 

0.6 

0.5 

0.6 

0.7 

6.9  16.60.6 

1.0 

1.0 

6 

\2kT.rogeTi... 

1.0 

0.0 

0.(! 

0. 0  0. 1 

0.3 

0.5 

0.1 

0.3 

0.4 

0.5 

0.3 

0.5 

0.6 

0.8  10.40.8 

1.0 

1.0 

7 

claviforme 

1.0 

0.0 

0.0 

0.20.3 

0.4 

3.6 

0.1 

0.3 

0.4 

0.6 

0.3 

0.6 

0.8 

0.9   0.60.6 

0.8 

1.0 

8 

lilacinum 

1.0 

+  1.0 

0.0 

0.0  0.0 

0.0 

;).o 

0.0 

D.O 

1).0 

0.0 

0.0 

0.1? 

0.1? 

0.1  10.0  0.3 

0.6 

0.6 

9 

granulatum 

1.0 

0.0 

0.0 

0.2  0.3 

0.4 

0.6 

0.3 

0.5 

0.6 

0.8 

0.3 

0.5 

0.6 

0.8  !0.60.8 

1.0 

1.0 

10 

italicum 

1.0 

0.0 

1.0 

0.1,0.8 

0.9 

0.9 

0.3 

0.7 

0.8 

0.9 

0.5 

0.8 

0.9 

1.0?  0.81.0 

1.0 

1.0 

11 

luteum 

1.0 

+  1.0 

0.0 

0. 0,0. 0 

0.1? 

0.1? 

0.0 

0.0 

0.0 

0.0 

0.0 

0.1? 

0.1? 

0.1?  0.1  0.3 

0.3 

O.S 

12 

1.0 

0.0 

1.0 

0.0 

0.0 

0.0 

0.1? 

0.0 

0.1? 

0. 1? 

0. 1? 

0.0 

0.0 

0.0 

0.0  0.20.2 

0.2 

0.2 

13 

1.0 

0.0 

0.00.2 

0.5 

0.6 

0.8 

0.3 

0.5 

0.7 

0.9 

0.4 

0.7 

0.9 

1.0 

0.6  1.0 

1.0 

1.0 

14 

expansum 

1.0 

0.0 

0.0 

0.1 

0.1 

0.3 

0.4 

0.2 

0.2 

0.7 

0.8 

0.3 

0.6 

0.6 

0.6 

0.6  0.9 

1.0 

1.0 

15 

citrinum 

1.0 

0.3 

0.0 

0.1 

0.0 

[).0 

;).  1? 

0.1 

0.  1? 

0. 1? 

0.  1? 

0.1 

0.1 

0.1 

0.1 

0.20.3 

0.3 

0.3 

16 

digitatum 

1.0 

0.0 

1.0 

0.1 

0.1 

0.3 

0.4 

0.2 

0.2 

0.3 

0.4 

0.1 

0.4 

0.5 

0  6 

0.5  0.7 

0.8 

0.9 

17 

purpurogenum 

1.0 

1.0 

0.0 

0.0 

0. 1? 

0.1? 

0. 1? 

0.1 

0.1 

0.1 

0.1 

0.0 

0. 1? 

0.1? 

0. 1? 

0.3  0.1 

0.1 

0.1 

18 

Toqueforli 

1.0 

0.0 

0.0 

0.2 

0.3 

:).7 

:).8 

0. 25 

0.5 

0. ; 

0.9 

0.4 

0.7 

1.0 

1.0 

0.61.0 

1.0 

1.0 

19 

roseum 

1.0 

0.0 

1.0 

0.0 

0.1 

XI 

;).  1 

0.0 

0.1 

0.1 

0.1 

0.1? 

0.0 

0.0 

0.0 

0.1  jo.  5 

0.5 

0.6 

20 

duclauzi 

1.0 

0.0 

0.0 

0.0 

0.0 

0.1 

0.1 

0.0 

0.0 

0.1 

0.1 

0.1? 

0.1? 

0.1 

0.1 

0.3 

0.4 

0.4 

?1 

1  0 

0  0 

0  0 

1  0 

)  0 

0  0 

0  0 

0  0 

0  0 

79 

1.0 
1.0 

0.0 
0.0 

0.0 
0.0 

23 

commune 

0.3 

0.6 

0.6 

0.8 

0.4 

0.6 

0.7 

0.9 

0.6 

0.9 

1.0 

1.0 

0.60.8 

1.0 

1.0 

24 

1.0 

0.0 

0.0 

0.1 

0.3 

3.5 

X7 

0.2 

0.5 

0.7 

0.9 

0.4 

0.7 

1.0 

1.0 

0.6  1.0 

1.0 

1.0 

?5 

—26 

1.0 
1.0 

0.8 
1.0 

0.0 
0.0 

26 

chrysogenum... 

0.1 

0.1 

0.3 

0.6 

0.4 

0.5 

0.6 

0.8 

0.4 

0.6 

0.8 

1.0 

6.6i.6 

1.0 

1.0 

27 

stoloniferum . . . 

1.0 

0.0 

1.0 

0.2 

0.1 

XI 

0.3? 

0.4 

0.4 

0.4 

0.5 

0.5 

0.5 

0.5 

0.6 

0.6 

0.9 

1.0 

1.0 

28 

1.0 

0.0 

1.0 

0.0 

0.0 

XI 

[).  1 

0.0 

0. 1? 

0.1 

0.3 

0.1 

0.3 

0.4 

0.6 

0.4 

0.6 

0.8 

0.9 

29 

1.0 

0.0 

0.0 

0.1 

0.3 

0.0 

0.1 

0.2 

0.6 

0.1 

0.5 

0.5 

0.6 

0.4 

0.7 

0.8 

0.9 

30 

1.0 

0.0 

0.0 

0.0 

0.1 

0.4 

0.5 

0.0 

0.0? 

(?) 

(?) 

0.0 

0.4 

0.7 

0.8 

0.0 

0.7? 

(?) 

(?) 

31 

1.0 

0.0 

0.0 

0.2 

0.5 

0.6 

0.6 

0.4 

0.5 

0.7 

0.8 

0.5 

0.8 

1.0 

1.0 

0.6 

0.7 

0.8 

0.9 

y> 

1  0 

+  1  0 

0  0 

33 

1.0 
1.0 

0.0 
+  1.0 

1.0 
0.0 

34 

divaricatum 

0.0 

0.0 

0.0 

0.0 

0.1? 

0.1 

0.1 

0.1 

0.1 

0.0 

0.0 

0.0 

0.1 

0.2 

0.3 

0.3 

35 

cf.  26 

1.0 
1.0 

36 

decumbens 

0.2 

0.0 

0.0 

0.0 

0.1 

0.2 

0. 2  '0. 1 

0.3 

0.4 

0.4 

0.2 

0.7 

0.7 

0.8 

37 

1.0 
1.0 

0.0 
0.0 

l.O'O.l 

0  y> 

0.1? 
0.3 

0.1? 
0.5 

0.1 
0.1 

0.1? 
0.1 

0.1? 
0.4 

0.1?'.... 
0.7   0.2 

1.0' 

1.6' 

1.6 

0.1 
0.8 

0.3 
0.9 

0.3 
1.0 

0.3 

38 

atramentosum . 

0.00.10.1 

1.0 

39 

biforme 

cf  23 

1.0 

0.0 

0.0  0.4,0.4 

0.7 

0.9 

0.4 

0.2 

0.5 

0.8  0.5 

0.6 

0.7 

0.9 

0.6 

0.9 

1.0 

1.0 

40 

1.0 
1.0 
1.0 

0.0 
0.0 
1.0 

0.0 
1.0 
0.0 

! 

41 

—  19 

...1.... 

1 

42 

funiculosum. .. 

0.0.0.0 

0.0 

0.1? 

0.0 

0.0 

0.1 

o.i  6.6 

0.3 

0.5 

0.7 

0.1 

0.7 

0.8 

0.9 

43 

cf.  17 

1.0 
1.0 
1.0 

1.0 
-0.4 
+  1.0 

0.00.00.0 

0.0 

0.1? 

0.1 

0.0 

0.1? 

0. 1?  0. 1 

0.1 

0.3 

0.4 

0.5 

44 

cf.  26 

0.0 
0.0 

45 

spinulosum 

6.60.  i? 

0.1 

0.3 

0.1 

0.1 

0.2 

0.3  10.4 

0.5 

0.7 

0.7 

6.6:0.7 

1.0 

1.0 

46 

Tugviosum 

1.0 

0.0 

0.00.20.1 

0.1 

0.2 

0.2 

0.1 

0.2 

0.3  I0.4 

0.5 

0.5 

0.6 

0.5  0.7 

0.8 

1.0 

47 
49 

1.0 
1.0 

0.3 
+  1.0 

0.0 
0.0 

intricatum 

0.00.0 

0.0 

0.1? 

0.0 

0.0 

0. 0  0. 1 

0.3 

0.3 

0.3 

6.i!6.3 

0.6 

0.6 

51 
54 
56 

1.0 
1.0 
1.0 

0.0 
0.0 
0.0 

0. 0  0. 1  0. 1 

0.1 

0.1 

0.0 

0.0 

0.0  .... 

0.3 

0.4 

0.4 

(?)0.7? 

0.8? 

1.0? 



0.0 
0.0 

—38 

1 

Explanation  of  decimals:  0.1  denotes  germination  of  conidia  only;  decimals  up  to  0.7,  growth  without 
the  formation  of  colored  conidial  areas;  0.7  to  1.0,  colonies  with  colored  conidia;  1.0  denotes  typical  colony. 
+  1.0  denotes  growth  more  rapid  at  37°  than  at  20°  C. 


106  CULTURAL  STUDIES  OF   SPECIES  OF  PENICILLIUM. 

REFERENCES  TO  LITERATURE. 

1.  Bessey,  E.  a.     Spore  forms  of  Spegazzinia  omata  Sacc.    Journal  of  Mycology, 

13,  43-45,  1907. 

2.  Brefeld,  Oscar.     Botanische  Untersuchungen  fiber  Schimmelpilze.     Heft  2: 

Die  Entwicklungsgeschichte  von  Penidllium.    Plates.    98  pp.,  Leipzig,  1874. 

3.  Clements,  Frederic  E.    Anecologic  view  of  the  species  conception.     Botanical 

Society  of  America.     Publication  34,  pp.  253-264,  Baltimore,  May  30, 1908. 

4.  Conn,  Herbert  W.     Bacteria  in  milk  and  its  products.     Illustrated,  306  pp., 

Philadelphia,  1903.     See  p.  268. 

5.  DiERCKx;  R.  P.     Un  essai  de  revision  du  genere  Penidllium  Link.    Annales  de  la 

Soci6t6  Scientifique  de  Bruxelles,  annexe  25,  1900-1901,  Ist  fascicule,  pp.  83-89, 
Lou  vain,  1901. 

6.  Eustace,  H.  J.     Investigations  on  some  fruit  diseases.     I.  Apple  rots  in  cold 

storage.  New  York  Agricultural  Experiment  Station,  Bulletin  297,  Geneva, 
1908. 

7.  Fries,  Elias.     Systema  mycologicum.    Vol.  3,  Gryphiswaldse,  1829.     See  p.  407- 

8.  Grevflle,  Robert  K.     Scottish  cryptogamic  flora,  or  colored  figures  and  descrip. 

tions  of  cryptogamic  plants,  belonging  chiefly  to  the  order  fungi.  Vols.  1-7, 
Edinburgh,  1823-1828. 

9.  Hedgcock,  George  G.,  and  Spauldinq,  Perley.    A  new  method  of  mounting 

fungi  grown  in  cultures  for  the  herbarium.  Journal  of  Mycology,  vol.  12,  No. 
84,  p.  147,  Columbus,  1906. 

10.  LiNDAU,  G.     Deutschl.  Krypt.  Flora.  Pilze,  vol.  8,  p.  166. 

11.  Link,  H.  F.     Linne  species  plantarum.     Editio  quarta,  tomus  6,  p.  70. 

12.  Observationes  in  ordines  plantarum  naturales.    Magazin  fiir  die  Neusten  Ent- 

deckungen  in  der  Gesammten  Naturkunde,  Jahrg.  3,  p.  16-18,  1809. 

13.  Maze,  P.     Les  microbes  dans  I'industrie  fromagdre.     Annales  de  I'lnstitut  Pas- 

teur, tome  19,  No.  6,  pp.  378^03,  June  25;  No.  8,  pp.  481-493,  August  25,  Paris, 
1905. 

14.  Micheli,   Petro  A.     Nova  plantarum  genera.     Illustrated,  234  pp.,  Florence, 

1729.     See  plate  91,  figures  1-4. 

15.  MiLBURN,  Thomas.     Ueber  Aenderungen  der  Farben  bei  Pilzen  und  Bakterien. 

Centralblatt  fiir  Bakteriologie,  abt  2,  band  13,  No.  5/7,  pp.  129-138,  Oct.  7;  No. 
9/11,  pp.  257-276,  Oct.  21,  Jena,  1904. 

16.  MoRiNi,  Fausto.     Sulla  forma  ascofora  del  Penidllium  candidum  Link.    Mal- 

pighia,  anno  2,  fascicule  5/6,  pp.  224-234,  Messina,  1888. 

17.  OuDEMANS,  C.  A.  J.  A.,  and  Koning,  C.  J.     Prodome  d'une  flore  mycologique 

obtenue  par  la  culture  sur  gelatine  prepar^e  de  la  terre  humeuse  du  Spanders- 
woud,  pr^  de  Bussum.  Archives  Neerlandaises  des  Sciences  Exactes  et 
Naturelles,  ser.  2,  tome  7,  No.  2/3,  pp.  266-298.     La  Haye,  1902.      See  p.  288. 

18.  Peck,  Charles  H.    Penidllium  pallidofulvum,  n.  sp.     New  York  State  Museum, 

Bulletin  67,  Report  of  the  State  Botanist  for  1902,  p.  30,  Albany,  1903. 

19.  Persoon,  D.  C.  H.     Synopsis  methodica  fungorum.    Part  2,  p.  693.     Gottingen, 

1801. 

20.  Powell,  G.  Harold,  Stubenrauch,  A.  V.,  Tenny,  L.  S.,  Eustace,  H.  J., 

HosFORD,  G.  W.,  and  White,  H.  M.  The  decay  of  oranges  while  in  transit  from 
California.  U.  S.  Department  of  Agriculture,  Bureau  of  Plant  Industry,  Bulle- 
tin 123,  Washington,  1908. 

21.  Roger,  Georges.     (Article  in)  Revue  Hebdomadaire,  vol.  7,  p.  334,  Paris. 

22.  Smith,  Ralph  E.    The  "soft  spot"  of  oranges.     {Penidllium  digitatum.)    Botan- 

ical Gazette,  vol.  24,  No.  2,  pp.  103-104,  Chicago,  1897. 

23.  California  Agricultural  Experiment  Station,  Bulletin  184.     Report  of  the  Plant 

Pathologist  to  July  1,  1906.     Sacramento,  1907. 


REFERENCES   TO  LITERATURE.  107 

24.  Stoll,  Otto.    Beitrage  zur  morphologischen  und  biologischen  Charakteristik  von 

Penicilliumarten.     Inaugural-Dissertation.     Plates,   56  pp.,  Wurzburg,  1904. 

25.  Thom,  Charles.     Fungi  in  cheese  ripening:  Camembert  and  Roquefort.     U.  S. 

Department  of  Agriculture,  Bureau  of  Animal  Industry,  Bulletin  82,  Wash- 
ington, 1906. 

26.  Storrs  Agricultural  Experiment  Station,  17th  Annual  Report,  1905,  pp.  73-115, 

1906. 

27.  Some  suggestions  from  the  study  of  dairy  fungi.    Journal  of  Mycology,  vol. 

11,  No.  77,  pp.  117-124,  Columbus,  1905. 

28.  Wehmer,  Carl.     Beitrage  zur  Kenntnis  einheimischer  Pilze.    Heft  1:  Zweineue 

Schimmelpilze  als  Erreger  einer  Citronensaure-garung.     Plates,  92  pp.,  Han- 
over and  Leipzig,  1893. 

29.  Zur  morphologie  und  entwickelungsgeschichte  des  Penidllium  luteum  Zuk., 

eines  iiberaus  haufigen  griinen  Schimmelpilzes.     Berichte  der  Deutschen 
Botanischen  Gesellschaft,  Band  11,  Heft  8,  pp.  499-516,  tafel  25,  Berlin,  1893. 

30.  Eine  neue  sklerotien-bildende  Penicillium-species  (P.  italicum).     Iledwigia. 

Organ  fiir  Kryptogamenkunde,  Band  33,  Heft  4,  pp.  211-214,  Dresden,  1894. 

31.  Beitrage  zur  Kenntnis  einheimischer  Pilze.     Heft  2:  Untersuchungen   iiber 

die  Faulnis  der  Friichte.     Plates,  84  pp.,  Jena,  1895.     See  p.  68,  plate  2. 

32.  Kleinere  mykologische  Mitteilungen.     Centralblatt  fiir  Bakteriologie,  abt.  2, 

Band  3,  No.  6,  pp.  147-153,  Jena,  Mar.  20,  1897.     See  p.  149. 

33 .  Ueber  die  Lebgnsdauer  eingetrockneter  Pilzkulturen .    Berichte  der  Deutschen 

Botanischen  Gesellschaft,  Band  22,  Heft  8,  pp.  476-478,  Berlin,  1904. 

34.  Morphologie,  Physiologic  und  Systematik  einiger  technisch  wichtiger  hoherer 

Ascomyceten  und  verwandter  Formen.     Lafar's  Handbuch  der  Technisohe 
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35.  Weidemann,  Carl.     Morphologische  und  physiologische  Beschreibung  einiger 

Penicillium-arten.     Centralblatt  fiir  Bacteriologie,  Abt.  2,  Band  19,  No.  21-23, 
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36.  Weigmann,  II.     Die  Giirungen  der  Milch  und  der  Abbau  ihrer  Bestandteile.     La- 

far's Handbuch  der  Technischen  Mykologie,  JBand  2,  Lieferung  9  and  12,  pp. 
48-189,  Jena,  1905-1906.     See  p.  186. 


INDEX  OF  SPECIES. 

Penidllium —  Page. 

album  Epstein 50 

atramentosum 65 

aureum  Corda,  emended  Hedgcock S7 

brevicaule  Saccardo 45 

brevicaule  Saccardo,  var.  album  n.  var 47 

brevicaule  Saccardo,  var.  glabrum  n.  var 48 

camemberti  Thorn 50 

camemberti  Thom,  var.  rogeri  n.  var 52 

candidum  Link 52 

chrysogenum  n.  sp 58 

citrinum  n.  sp 61 

claviforme  Bainier 43 

commune  n.  sp 56 

decumbens  n.  sp 71 

digitatum  Saccardo 31 

divaricatum  n.  sp 72 

duclauxi  Delacroix 42 

epsteinii  Lindau 50 

expansum  Link,  emended 27 

funiculosum  n.  sp 69 

glaucum  Link 26 

granulatum  Bainier 44 

intricatum  n.  sp 75 

italicum  Wehmer 29 

liladnum  n .  sp 73 

luteum  Zukal 39 

olivaceum  Wehmer 31 

pinopMlum  Hedgcock,  n.  nov 37 

purpurogenum  O.  Stoll 36 

roqueforti  Thom 34 

roseum  Link  (?) 49 

rubrum  O.  Stoll 39 

rugulosum  n.  sp 60 

spinulosum  n.  sp 76 

stoloniferum  n.  sp 68 

109 

o 


